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Sample records for human granulosa lutein

  1. Establishment and validation of a model for non-luteinized human mural granulosa cell culture.

    Science.gov (United States)

    Ophir, L; Yung, Y; Maman, E; Rubinstein, N; Yerushalmi, G M; Haas, J; Barzilay, E; Hourvitz, A

    2014-03-25

    Cell culture techniques of human mural granulosa cells (MGCs) serve as a major in vitro tool. However, the use of luteinized MGCs has major limitations due to their luteinized state. Our aim was to establish a standardized protocol for the culture of MGCs as a model for different stages of folliculogenesis. We showed that early-non-luteinized, preovulatory-non-luteinized and luteal-MGCs have distinct gene expression pattern. After 4 days of incubation of luteinized-MGCs, ovulatory genes mRNA's achieve expression levels similar to the early non-luteinized follicles. FSH stimulation for 48 h of these 4 days cultured MGCs showed ovulatory genes mRNA's expression similar to the pre-ovulatory non-luteinized follicles. These FSH-stimulated cells responded to hCG stimulation in a pattern similar to the response of pre-ovulatory follicles. This novel model may provide a standardized research tool for delineation of the molecular processes occurring during the latter stages of follicular development in the human ovary.

  2. Potential role of hCG in apoptosis of human luteinized granulosa cells

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    HIRATA, Rei; HOJO, Takuo; SANO, Masahiro; HAYASHI, Nobuyoshi; OKUDA, Kiyoshi

    2014-01-01

    The corpus luteum (CL) forms after ovulation and acts as a temporary endocrine gland that produces progesterone (P4), a hormone that is essential for implantation and maintenance of pregnancy in mammals. In pregnant women, human chorionic gonadotropin (hCG) secreted by the conceptus prevents luteolysis. hCG also increases the survival of cultured human luteinized granulosa cells (hLGCs). To clarify the maintenance mechanism of the human CL, we investigated the effects of hCG and P4 receptor antagonists, onapristone (OP) and RU486, on the viability of hLGCs. With the patients’ consent, hLGCs were isolated from follicular aspirates for in vitro fertilization. The cells were cultured with hCG (0.1, 1, 10, 100 IU/ml), OP (10, 25, 50, 100 μM), RU486 (100 μM), P4 (1, 10, 25, 50 μM) or some combination of the four for 24 h. Cell viability was significantly increased by hCG (100 IU/ml) and significantly decreased by OP (100 μM) compared with the control. Cells treated with hCG and OP together were significantly less viable than the control and OP-treated cells. The combined treatment also significantly increased CASP3 activity and cleaved CASP3 protein expression. Furthermore, P4 addition reversed the reduction in cell viability caused by the combination of hCG and OP treatment. The overall findings suggest that hCG cooperates with P4 to increase survival of hLGCs and to induce apoptosis when P4 action supported by hCG is attenuated in the human CL. PMID:25451535

  3. Vasoactive intestinal polypeptide and peptide histidine methionine. Presence in human follicular fluid and effects on DNA synthesis and steroid secretion in cultured human granulosa/lutein cells

    DEFF Research Database (Denmark)

    Gräs, S; Ovesen, P; Andersen, A N;

    1994-01-01

    Vasoactive intestinal polypeptide (VIP) and peptide histidine methionine (PHM) originate from the same precursor molecule, prepro VIP. In the present study we examined the concentrations of VIP and PHM in human follicular fluid and their effects on cultured human granulosa/lutein cells. Follicular...

  4. Vasoactive intestinal polypeptide and peptide histidine methionine. Presence in human follicular fluid and effects on DNA synthesis and steroid secretion in cultured human granulosa/lutein cells

    DEFF Research Database (Denmark)

    Gräs, S; Ovesen, P; Andersen, A N;

    1994-01-01

    fluid and cells were obtained from patients undergoing in-vitro fertilization for tubal infertility. The concentrations of VIP and PHM in pre-ovulatory human follicular fluid were measured radioimmunochemically. Granulosa/lutein cells isolated from follicular fluid were cultured under serum......Vasoactive intestinal polypeptide (VIP) and peptide histidine methionine (PHM) originate from the same precursor molecule, prepro VIP. In the present study we examined the concentrations of VIP and PHM in human follicular fluid and their effects on cultured human granulosa/lutein cells. Follicular......-free conditions with VIP and PHM in varying concentrations (0.1, 10, 1000 nmol/l). [3H]Thymidine incorporation in the cells and oestradiol as well as progesterone concentrations in the culture medium were measured. The mean (+/- SEM) concentrations of VIP and PHM were 6.8 +/- 0.1 and 7.7 +/- 0.8 pmol...

  5. Bisphenol-A exposure and gene expression in human luteinized membrana granulosa cells in vitro.

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    Mansur, Abdallah; Israel, Ariel; Combelles, Catherine M H; Adir, Michal; Racowsky, Catherine; Hauser, Russ; Baccarelli, Andrea A; Machtinger, Ronit

    2017-02-01

    Does bisphenol-A (BPA) affect gene expression in human membrana granulosa cells (MGC)? In vitro, short exposure to supra-physiological concentrations of BPA alters human MGC gene expression. Exposure to BPA may interfere with reproductive endocrine signaling. In vitro studies, mostly in animal models, have shown an inverse correlation between exposure to BPA and follicular growth, meiosis, and steroid hormone production in granulosa cells. Primary cultures of MGC obtained from 24 patients undergoing IVF (for PGD, male factor infertility or unexplained infertility) were exposed to various concentrations of BPA (0, 0.02, 0.2, 2 or 20 µg/ml) for 48 h. The study was conducted in a university-affiliated hospital. Microarray analysis was used to identify genes exhibiting expression changes following BPA exposure. Genes significantly altered were identified based on changes greater than 2-fold relative to the control group (not treated by BPA) and a Student's t-test P-value <0.05. Statistical significance was adjusted for multiple comparisons using the Benjamini-Hochberg method. Alterations in the expression of genes that are involved in the enriched functional annotations altered by BPA at the concentration of 20 µg/ml were confirmed by real-time PCR. A distinct pattern of gene expression was observed in primary cultures of MGC exposed to the highest BPA concentration compared with untreated cells. We identified 652 genes that exhibited at least 2-fold differences in expression after BPA exposure (all P < 0.05 versus untreated). These genes were significantly enriched for annotations related to cell cycle progression, segregation of chromosomes, steroid metabolism, apoptosis, lipid synthesis, oocyte maturation and chromosomal alignment. No significant changes in gene expression were found at the lower doses of BPA most relevant to human exposure. N/A. Human exposure to BPA in vivo occurs over long periods of time. In this in vitro model, cells were exposed to the

  6. Follicle-stimulating hormone potentiates the steroidogenic activity of chorionic gonadotropin and the anti-apoptotic activity of luteinizing hormone in human granulosa-lutein cells in vitro.

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    Casarini, Livio; Riccetti, Laura; De Pascali, Francesco; Nicoli, Alessia; Tagliavini, Simonetta; Trenti, Tommaso; La Sala, Giovanni Battista; Simoni, Manuela

    2016-02-15

    Luteinizing hormone (LH) and choriogonadotropin (hCG) are glycoprotein hormones regulating ovarian function and pregnancy, respectively. Since these molecules act on the same receptor (LHCGR), they were traditionally assumed as equivalent in assisted reproduction techniques (ART), although differences between LH and hCG were demonstrated at molecular and physiological level. In this study, we demonstrated for the first time that co-treatment with a follicle-stimulating hormone (FSH) dose in the ART therapeutic range potentiates different LH- and hCG-dependent responses in vitro, measured in terms of cAMP, phospho-CREB, -ERK1/2 and -AKT activation, gene expression, progesterone and estradiol production in human granulosa-lutein cells (hGLC). We show that in the presence of FSH, hCG biopotency is about 5-fold increased, in the presence of FSH, in terms of cAMP activation. Accordingly, CREB phosphorylation and steroid production is increased under hCG and FSH co-treatment. LH effects, evaluated as steroidogenic cAMP/PKA pathway activation, do not change in the presence of FSH, which, however, increases LH-dependent ERK1/2 and AKT, but not CREB phosphorylation, resulting in anti-apoptotic effects. The different modulatory activity of FSH on LH and hCG action in vitro corresponds to their different physiological functions, reflecting proliferative effects exerted by LH during the follicular phase and before trophoblast development, and the high steroidogenic potential of hCG requested to sustain pregnancy from the luteal phase onwards.

  7. Induction of Ski Protein Expression upon Luteinization in Rat Granulosa Cells.

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    Kim, Hyun; Kim, Dong Hun; Park, Soo Bong; Ko, Yeoung-Gyu; Kim, Sung-Woo; Do, Yoon Jun; Park, Jae-Hong; Yang, Boh-Suk

    2012-05-01

    Ski protein is implicated in proliferation/differentiation in a variety of cells. We had previously reported that Ski protein is present in granulosa cells of atretic follicles, but not in preovulatory follicles, suggesting that Ski has a role in apoptosis of granulosa cells. The alternative fate of granulosa cells other than apoptosis is to differentiate to luteal cells; however, it is unknown whether Ski is expressed and has a role in granulosa cells undergoing luteinization. Thus, the aim of the present study was to locate Ski protein in the rat ovary during luteinizationto predict the possible role of Ski. In order to examine the expression pattern of Ski protein along with the progress of luteinization, follicular growth was induced by administration of equine chorionic gonadtropin to immature female rats, and luteinization was induced by human chorionic gonadtropin treatment to mimic luteinizing hormone (LH) surge. While no Ski-positive granulosa cells were present in preovulatory follicle, Ski protein expression was induced in response to LH surge, and was maintained after the formation of the corpus luteum (CL). Though Ski protein is absent in granulosa cells of preovulatory follicle, its mRNA (c-Ski) was expressed and the level was unchanged even after LH surge. Taken together, these results demonstrated that Ski protein expression is induced in granulosa cells upon luteinization, and suggests that its expression is regulated post-transcriptionally.

  8. Induction of Ski Protein Expression upon Luteinization in Rat Granulosa Cells

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    Hyun Kim

    2012-05-01

    Full Text Available Ski protein is implicated in proliferation/differentiation in a variety of cells. We had previously reported that Ski protein is present in granulosa cells of atretic follicles, but not in preovulatory follicles, suggesting that Ski has a role in apoptosis of granulosa cells. The alternative fate of granulosa cells other than apoptosis is to differentiate to luteal cells; however, it is unknown whether Ski is expressed and has a role in granulosa cells undergoing luteinization. Thus, the aim of the present study was to locate Ski protein in the rat ovary during luteinizationto predict the possible role of Ski. In order to examine the expression pattern of Ski protein along with the progress of luteinization, follicular growth was induced by administration of equine chorionic gonadtropin to immature female rats, and luteinization was induced by human chorionic gonadtropin treatment to mimic luteinizing hormone (LH surge. While no Ski-positive granulosa cells were present in preovulatory follicle, Ski protein expression was induced in response to LH surge, and was maintained after the formation of the corpus luteum (CL. Though Ski protein is absent in granulosa cells of preovulatory follicle, its mRNA (c-Ski was expressed and the level was unchanged even after LH surge. Taken together, these results demonstrated that Ski protein expression is induced in granulosa cells upon luteinization, and suggests that its expression is regulated post-transcriptionally.

  9. Induction of Ski protein expression upon luteinization in rat granulosa cells without a change in its mRNA expression.

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    Kim, Hyun; Yamanouchi, Keitaro; Matsuwaki, Takashi; Nishihara, Masugi

    2012-01-01

    The Ski protein is implicated in the proliferation/differentiation of a variety of cells. We previously reported that the Ski protein is present in granulosa cells of atretic follicles, but not in preovulatory follicles, suggesting that Ski has a role in apoptosis of granulosa cells. However, granulosa cells cannot only undergo apoptosis but can alternatively differentiate into luteal cells. It is unknown whether Ski is expressed and has a role in granulosa cells undergoing luteinization. Thus, the aim of the present study was to determine the localization of the Ski protein in the rat ovary during luteinization to examine if Ski might play a role in this process. In order to examine the Ski protein expression during the progression of luteinization, follicular growth was induced in immature female rats by administration of equine chorionic gonadotropin, and luteinization was induced by human chorionic gonadotropin treatment to mimic the luteinizing hormone (LH) surge. While no Ski-positive granulosa cells were present in the preovulatory follicle, Ski protein expression was induced in response to the LH surge and was maintained after formation of the corpus luteum (CL). Although the Ski protein is absent from the granulosa cells of the preovulatory follicle, its mRNA (c-ski) was expressed, and the level of c-ski mRNA was unchanged even after the LH surge. The combined results demonstrated that Ski protein expression is induced in granulosa cells upon luteinization, and suggested that its expression is regulated posttranscriptionally.

  10. Expression of Leptin Long-form Receptor mRNA in Luteinized Granulosa Cells of Obese Women with Polycystic Ovary Syndrome

    Institute of Scientific and Technical Information of China (English)

    YIN Jie; LIU Yi; LV Liqun; WANG Donghua; GONG Cheng; XIAO Wei; SHENG Hui

    2007-01-01

    To investigate the expression of mRNA of leptin long-form receptor (OB-Rb) in luteinized granulosa cells of obese women with polycystic ovary syndrome (PCOS), and to determine the role of leptin in the physiopathology of PCOS, luteinized granulosa cells were collected from the follicle fluid of 10 obese women who met the diagnostic criteria for PCOS and their BMI was equal to or greater than 25 kg/m2, and at the same time, granulosa cells were collected from 10 normal women undergoing IVF-ET who served as the control group. Some luteinized granulosa cells were taken from normal women for in-vitro culture, into which human leptin of different concentrations was added (0, 10, 100 and 1000 ng/mL). After stimulation with leptin for 48 h, RT-PCR was employed for the detection of the expression of OB-RLmRNA in the luteinized granulosa cells. Our results showed that the level of OB-RLmRNA in luteinized granulosa cells of obese PCOS women was higher than those in the control (P<0.05). In luteinized granulosa cells cultured in vitro and stimulated by human leptin for 48 h, the level of OB-RLmRNA was higher than those without leptin stimulation (P<0.01), and when leptin concentration was at 100 ng/mL, and the level of OB-RLmRNA reached a peak. It is concluded that in obese PCOS women, the level of serum leptin is increased, which promotes the expression of OB-RL in luteinized granulosa cells and increases the sensitivity of the granulosa cells to leptin. Leptin may contribute to anovulation in obese women with PCOS.

  11. Very low-dose (femtomolar) 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) disrupts steroidogenic enzyme mRNAs and steroid secretion by human luteinizing granulosa cells.

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    Baldridge, M G; Marks, G T; Rawlins, R G; Hutz, R J

    2015-04-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is the most toxic congener of the polyhalogenated aromatic hydrocarbons (PAH), which causes anatomical abnormalities and developmental defects, impairs ovulation and reduces fertility. TCDD's endocrine-disrupting effects are, in part, caused by a direct action at the ovary. Herein we investigated the in-vitro effects of environmentally relevant doses of TCDD on estradiol-17β (E2) production by human luteinizing granulosa cells (hLGC) obtained from women stimulated for in-vitro fertilization (IVF). TCDD at all concentrations tested (3.1fM, 3.1pM and 3.1nM) significantly decreased E2 secretion when assayed for by radioimmunoassay (RIA). Herein we confirm that TCDD alters E2 secretion by hLGC in a time-, not dose-dependent fashion and are the first to show decreases in E2 secretion with fM concentrations of TCDD. Using real-time quantitative PCR (RT-qPCR), the decreased E2 secretion correlates with a decrease in the mRNA expression levels two enzymes in the estrogen biosynthesis pathway: CYP11A1 and CYP19A1.

  12. 电压门控钾离子通道对卵巢黄素化颗粒细胞增殖、分泌及凋亡的影响%Influence of 4-aminopyridine on human ovarian luteinized granulosa cell proliferation, production, and apoptosis through inhibiting voltage-gated K+ channel

    Institute of Scientific and Technical Information of China (English)

    赵志明; 崔娜; 徐素欣; 高福禄; 郝桂敏; 曹金凤

    2008-01-01

    Objective To study the influence of 4-aminopyridine(4-AP)on proliferation,production,and apoptosis through inhibiting voltage-gated K+channel(Kv)in ovarian luteinized granulosa cells.nethods Ovarian luteinized granulosa cells were recovered from 25 women with regular menses who underwent in vitro fertilization programme.The cultured granulosa cells were divided into 4 groups:blank group,4-AP treated group,human chorionic gonadotropin(hCG)-induced group and hCG+4-AP cotreated group.The final concentrations of hCG and 4-AP were 1250 U/L and 5 nmol/L respectively.The progesterone production WaS detected by the chemoluminescence method.The expression of Kv mRNA on human ovarian luteinized granulosa cell was detected by RT.PCR The influence on the early apoptosis of gTanulosa cells bv 4-AP was observed by flow eytometry.Cellular caSpage-3 activities were observed with colorimetric method and the inhibition of the cell proliferation was studied using methyl thiazolyl tetrazolium(MTT)method.Results(1)Kv mRNA wag expressed in granulosa cell.(2)The progesterone production64),(206±32),(1991±172)and(763±79)nmol/L,respectively after24 hours culture.Exposure of the(3)The flow cytometry analysis and the cellular caapase-3 A405 showed that 4-AP increased the percentage ofearly phase apoptosis(P0.05).(3)培养24 h后颗粒细胞的凋亡率及半胱氨酸天冬氨酸蛋白酶(caspase-3)活性,4-AP组分别为(40±5)%和0.049 ±0.009,均高于空白组[(17±4)%和0.029±0.008],hCG+4-AP组[(25±4)%和0.039±0.008]也均高于hCG组[(15 ±3)%和0.022 ±0.007],差异均有统计学意义(P<0.01).结论 4-AP能够抑制卵巢黄素化颗粒细胞和经hCG诱导的颗粒细胞的孕酮分泌,町能与其抑制增殖、促进凋亡有关.Kv在卵巢黄素化颗粒细胞分泌、增殖及凋亡过程中起重要的作用.

  13. 瘦素对人卵巢黄素化颗粒细胞雌、孕激素分泌的体外实验研究%Effects of leptin on estradiol and progesterone production by human luteinized granulosa cells in vitro

    Institute of Scientific and Technical Information of China (English)

    郭新宇; 陈士岭; 邢福祺

    2001-01-01

    目的探讨瘦素在体外对人卵巢颗粒细胞雌激素和孕激素生成的影响。方法将来自体外受精-胚胎移植(IVF-ET)的卵巢黄素化颗粒细胞纯化后,在不同浓度瘦素(0、10、30、100、300 ng/ml) 和人绝经期促性腺激素(hMG,0、0.1、0.2、0.5、1、2、5、10 IU/ml)单独或联合作用下进行体外培养,收集培养液,采用放射免疫方法测定颗粒细胞产生雌二醇及孕酮的量。结果不同浓度的瘦素对雌二醇、孕酮的生成无影响;hMG为5 IU/ml时,雌二醇水平平均为2.36×10-11 mol/L,加入不同浓度瘦素(10、30、100、300 ng/ml),雌二醇的生成均受到明显的抑制,随瘦素浓度增加,雌二醇水平逐渐降低(P0.05) by human luteinized granulosa cells. Leptin of 10~30 ng/ml concentrations caused a dose-dependent inhibition of estradiol production (P0.05). Conclusion Leptin can directly inhibit hMG-stimulated estradiol production by human luteinized granulosa cells in vitro, but has no effect on progesterone production. Leptin may play an important role in follicle development and luteinization.

  14. Autocrine role of estrogens in the augmentation of luteinizing hormone receptor formation in cultured rat granulosa cells.

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    Kessel, B; Liu, Y X; Jia, X C; Hsueh, A J

    1985-06-01

    The effects of estrogens on gonadotropin-stimulated luteinizing hormone (LH) receptor formation were examined in primary cultures of rat granulosa cells. Granulosa cells were cultured for 3 days with increasing concentrations of follicle-stimulating hormone (FSH) in the presence or absence of native and synthetic estrogens. Follicle-stimulating hormone stimulated LH receptor formation in a dose-dependent fashion, and estrogens enhanced the FSH-stimulated LH receptor content by decreasing the apparent ED50 of FSH. At 6.25 ng/ml FSH, the enhancement in LH receptor was estrogen dose dependent, with an ED50 value of about 3 X 10(-9) M for 17 beta-estradiol. The increased LH receptor content seen in cells treated with FSH and estrogen was correlated with increased cAMP production by these cells in response to LH stimulation. Time course studies revealed enhancement of FSH-stimulated LH receptor induction at 48 and 72 h of culture. Granulosa cells were also cultured with FSH for 2 days to induce functional LH receptors, then further cultured for 3 days with LH in the presence or absence of estrogens. At 30 ng/ml LH, increasing concentrations of estrogens maintained LH receptor content in a dose-dependent fashion, with their relative estrogenic potencies in keeping with reported binding affinities to estrogen receptors. An autocrine role of estrogens on LH receptor formation was further tested in granulosa cells treated with FSH and an aromatase substrate (androstenedione) to increase estrogen biosynthesis. Cotreatment with semipurified estrogen antibodies partially blocked the FSH stimulation of LH receptors, whereas nonimmune serum was ineffective. Also, inclusion of diethylstilbestrol prevented the inhibitory effect of the estrogen antibodies. Thus, local estrogens in ovarian follicles may play an autocrine role in granulosa cells to enhance LH receptor formation and to increase granulosa cell responsiveness to the LH surge, with subsequent ovulation and adequate

  15. Luteinizing Hormone-Induced RUNX1 Regulates the Expression of Genes in Granulosa Cells of Rat Periovulatory Follicles

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    Jo, Misung; Curry, Thomas E.

    2006-01-01

    The LH surge induces specific transcription factors that regulate the expression of a myriad of genes in periovulatory follicles to bring about ovulation and luteinization. The present study determined 1) the localization of RUNX1, a nuclear transcription factor, 2) regulation of Runx1 mRNA expression, and 3) its potential function in rat ovaries. Up-regulation of mRNA and protein for RUNX1 is detected in preovulatory follicles after human chorionic gonadotropin (hCG) injection in gonadotropin-treated immature rats as well as after the LH surge in cycling animals by in situ hybridization and immunohistochemical and Western blot analyses. The regulation of Runx1 mRNA expression was investigated in vitro using granulosa cells from rat pre-ovulatory ovaries. Treatments with hCG, forskolin, or phorbol 12 myristate 13-acetate stimulated Runx1 mRNA expression. The effects of hCG were reduced by inhibitors of protein kinase A, MAPK kinase, or p38 kinase, indicating that Runx1 expression is regulated by the LH-initiated activation of these signaling mediators. In addition, hCG-induced Runx1 mRNA expression was inhibited by a progesterone receptor antagonist and an epidermal growth factor receptor tyrosine kinase inhibitor, whereas amphiregulin stimulated Runx1 mRNA expression, demonstrating that the expression is mediated by the activation of the progesterone receptor and epidermal growth factor receptor. Finally, knockdown of Runx1 mRNA by small interfering RNA decreased progesterone secretion and reduced levels of mRNA for Cyp11a1, Hapln1, Mt1a, and Rgc32. The hormonally regulated expression of Runx1 in periovulatory follicles, its involvement in progesterone production, and regulation of preovulatory gene expression suggest important roles of RUNX1 in the periovulatory process. PMID:16675540

  16. The role for runt related transcription factor 2 (RUNX2) as a transcriptional repressor in luteinizing granulosa cells.

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    Park, Eun-Sil; Park, Jiyeon; Franceschi, Renny T; Jo, Misung

    2012-10-15

    Transcription factors induced by the LH surge play a vital role in reprogramming the gene expression in periovulatory follicles. The present study investigated the role of RUNX2 transcription factor in regulating the expression of Runx1, Ptgs2, and Tnfaip6 using cultured granulosa cells isolated from PMSG-primed immature rats. hCG or forskolin+PMA induced the transient increase in Runx1, Ptgs2, and Tnfaip6 expression, while the expression of Runx2 continued to increase until 48 h. The knockdown of the agonist-stimulated Runx2 expression increased Runx1, Ptgs2, and Tnfaip6 expression and PGE(2) levels in luteinizing granulosa cells. Conversely, the over-expression of RUNX2 inhibited the expression of these genes and PGE(2) levels. The mutation of RUNX binding motifs in the Runx1 promoter enhanced transcriptional activity of the Runx1 promoter. The knockdown and overexpression of Runx2 increased and decreased Runx1 promoter activity, respectively. ChIP assays revealed the binding of RUNX2 in the Runx1 and Ptgs2 promoters. Together, these novel findings provide support for the role of RUNX2 in down-regulation of Runx1, Ptgs2, and Tnfaip6 during the late ovulatory period to support proper ovulation and/or luteinization.

  17. Expression of inhibin α, matrix metalloproteinase-2 and tissue inhibitors of metalloproteinase-1 in luteinized granulosa cells: comparison between women with and without endometriosis

    Institute of Scientific and Technical Information of China (English)

    Tan Rong-rong; Liu Li-peng; Pu Dan-hua; Cui Yu-gui; Jiang Shi-Wen; Liu Jia-yin; Wu Jie

    2012-01-01

    Objective: To assess the differences in expression of inhibin α,MMP-2 and TIMP-1 in luteinized granulosa cells from women with and without endometriosis undergoing in vitro fertilization (IVF),and determine whether inhibin α,MMP-2 and TIMP-1 expressions correlate with the follicle development in patients with endometriosis.Methods: Thirty six infertile patients with stage Ⅲ and Ⅳ endometriosis patients (group A) and 35 tubal infertile and/or male factor patients (group B) attending to the clinic of Jiangsu Provincial Hospital Human Reproductive Center during May 2011 and December 2011 were included in this prospective study.The patients with the number of mature oocyte ≤ 4 were considered as poor responders.According to this,the two groups were further divided into four subgroups: A1 group (23 endometriosis patients with normal response),A2 group (13 endometriosis patients with poor response),B1 group (23 controls with normal response) and B2 group (12 controls with poor response).Fasting blood samples were collected on the first day of ovarian stimulation and on the day of hCG administration,and follicle fluid were collected on the day of oocytes retrieved.The concentrations of follicle stimulating hormone (FSH),Luteinizing hormone (LH) and estradiol (E2) were measured by ECILA.Granulosa was collected from follicle fluid by Percoll method,and Stepone real-time polymerase chain reaction (PCR) was used to analysis the expression level of inhibin α,MMP-2 and TIMP-1.Results: The expression of inhibin α in endometriosis group and poor responders in control group were significantly lower than those of the normal responders in control group,in which the alternation of inhibin α expression was not correlated to the variables of follicle development.Moreover,MMP-2 levels were lower and TIMP-1 levels were higher in both endometriosis and tubal/male infertility poor responders compared to normal ovarian responders.Interestingly,the correlation between MMP-2 or

  18. Opiate receptor blockade on human granulosa cells inhibits VEGF release.

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    Lunger, Fabian; Vehmas, Anni P; Fürnrohr, Barbara G; Sopper, Sieghart; Wildt, Ludwig; Seeber, Beata

    2016-03-01

    The objectives of this study were to determine whether the main opioid receptor (OPRM1) is present on human granulosa cells and if exogenous opiates and their antagonists can influence granulosa cell vascular endothelial growth factor (VEGF) production via OPRM1. Granulosa cells were isolated from women undergoing oocyte retrieval for IVF. Complementary to the primary cells, experiments were conducted using COV434, a well-characterized human granulosa cell line. Identification and localization of opiate receptor subtypes was carried out using Western blot and flow cytometry. The effect of opiate antagonist on granulosa cell VEGF secretion was assessed by enzyme-linked immunosorbent assay. For the first time, the presence of OPRM1 on human granulosa cells is reported. Blocking of opiate signalling using naloxone, a specific OPRM1 antagonist, significantly reduced granulosa cell-derived VEGF levels in both COV434 and granulosa-luteal cells (P opiate receptors and opiate signalling in granulosa cells suggest a possible role in VEGF production. Targeting this signalling pathway could prove promising as a new clinical option in the prevention and treatment of ovarian hyperstimulation syndrome.

  19. Expression of scavenger receptor-BI and low-density lipoprotein receptor and differential use of lipoproteins to support early steroidogenesis in luteinizing macaque granulosa cells.

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    Cherian-Shaw, Mary; Puttabyatappa, Muraly; Greason, Erin; Rodriguez, Annabelle; VandeVoort, Catherine A; Chaffin, Charles L

    2009-02-01

    An ovulatory hCG stimulus to rhesus macaques undergoing controlled ovarian stimulation protocols results in a rapid and sustained increase in progesterone synthesis. The use of lipoproteins as a substrate for progesterone synthesis remains unclear, and the expression of lipoprotein receptors [very-low-density lipoprotein receptor (VLDLR), low-density lipoprotein receptor (LDLR), and scavenger receptor-BI (SR-BI)] soon after human chorionic gonadotropin (hCG) (lipoprotein receptor expression and lipoprotein (VLDL, LDL, and HDL) support of steroidogenesis during luteinization of macaque granulosa cells. Granulosa cells were aspirated from rhesus monkeys undergoing controlled ovarian stimulation before or up to 24 h after an ovulatory hCG stimulus. The expression of VLDLR decreased within 3 h of hCG, whereas LDLR and SR-BI increased at 3 and 12 h, respectively. Granulosa cells isolated before hCG were cultured for 24 h in the presence of FSH or FSH plus hCG with or without VLDL, LDL, or HDL. Progesterone levels increased in the presence of hCG regardless of lipoprotein addition, although LDL, but not HDL, further augmented hCG-induced progesterone. Other cells were cultured with FSH or FSH plus hCG without an exogenous source of lipoprotein for 24 h, followed by an additional 24 h culture with or without lipoproteins. Cells treated with hCG in the absence of any lipoprotein were unable to maintain progesterone levels through 48 h, whereas LDL (but not HDL) sustained progesterone synthesis. These data suggest that an ovulatory stimulus rapidly mobilizes stored cholesterol esters for use as a progesterone substrate and that as these are depleted, new cholesterol esters are obtained through an LDLR- and/or SR-BI-mediated mechanism.

  20. Dopamine receptor repertoire of human granulosa cells

    Directory of Open Access Journals (Sweden)

    Kunz Lars

    2007-10-01

    Full Text Available Abstract Background High levels of dopamine (DA were described in human ovary and recently evidence for DA receptors in granulosa and luteal cells has been provided, as well. However, neither the full repertoire of ovarian receptors for DA, nor their specific role, is established. Human granulosa cells (GCs derived from women undergoing in vitro fertilization (IVF are an adequate model for endocrine cells of the follicle and the corpus luteum and were therefore employed in an attempt to decipher their DA receptor repertoire and functionality. Methods Cells were obtained from patients undergoing IVF and examined using cDNA-array, RT-PCR, Western blotting and immunocytochemistry. In addition, calcium measurements (with FLUO-4 were employed. Expression of two DA receptors was also examined by in-situ hybridization in rat ovary. Effects of DA on cell viability and cell volume were studied by using an ATP assay and an electronic cell counter system. Results We found members of the two DA receptor families (D1- and D2 -like associated with different signaling pathways in human GCs, namely D1 (as expected and D5 (both are Gs coupled and linked to cAMP increase and D2, D4 (Gi/Gq coupled and linked to IP3/DAG. D3 was not found. The presence of the trophic hormone hCG (10 IU/ml in the culture medium for several days did not alter mRNA (semiquantitative RT-PCR or protein levels (immunocytochemistry/Western blotting of D1,2,4,5 DA receptors. Expression of prototype receptors for the two families, D1 and D2, was furthermore shown in rat granulosa and luteal cells by in situ hybridization. Among the DA receptors found in human GCs, D2 expression was marked both at mRNA and protein levels and it was therefore further studied. Results of additional RT-PCR and Western blots showed two splice variants (D2L, D2S. Irrespective of these variants, D2 proved to be functional, as DA raised intracellular calcium levels. This calcium mobilizing effect of DA was observed

  1. Effects of growth hormone on estradiol and progesterone production in human luteinized granulosa cells in vitro%生长激素对卵泡黄素化颗粒细胞分泌雌、孕激素的影响

    Institute of Scientific and Technical Information of China (English)

    曾晓霞; 刘风华; 李玲; 石宇; 杜红姿

    2012-01-01

    Objective To determine whether there were differences in the dose-dependent effects of growth hormone (GH) on estradiol(E2) ,progesterone(P) , insulin-like growth factor-1 (IGF-1) production by luteinized granulosa cells. To detect GH and IGF-1 receptors in human luteinized granulosa cells. Methods Human luteinized granulosa cells (LGC) were isolated from 24 women undergoing in vitro fertilization and embryo transfer (IVF-ET) in the Third Affiliated Hospital of Guangzhou Medical College between May and July 2010. Some of the LGCs were used for detecting GH and IGF-1 receptors by immunocytochemistry(SABC method). Most of them were cultured in DMEM/F12 medium with 10% fetal bovine serum(FCS) , In 2 days,the medium was removed and the cells were incubated with various concentration of GH (0,4. 7 ,9. 4 and 18. 8 nmol/L) in the presence or absence of follicle stimulating hormone(FSH 75IU/L). Media were collected in 72 hours. E2 ,P in media were measured with chemiluminescence immune assay (CLIA) , while IGF-1 was measured with enzyme linked immunosorbent assay (ELISA). Results More than 80% of the LGCs were positively stained for GH and IGF-1 receptors. LGCs could autocrine IGF-1 , E2, and P at the level of 140. 00 ± 27. 14 μg/L,444. 67 ±67. 25 pmol/L, 655. 00 ±70. 07 nmol/L without GH or FSH co-culture. Cultured with medium plus various concentration of GH (4. 7 ,9. 4,18. 8 nmol/L) , LGC biosyn-thesized concentration of IGF-1, E2, and P were (140. 33 ± 18. 60,139. 08 ±31.81,154.72 ±32. 19 μg/L) , (489.00 ± 91.27,658.00 ±219.67,660.83 ±207.95 pmol/L) and (821.67 ±101.27,995. 00 ±109. 68,1193. 31 ±138.80 nmol/L) respectively,and there was not significant difference, compared with basic state. Level of IGF-1 , E2 , and P were (149. 41 ± 50. 30 ,155. 34 ± 36. 34,154. 33 ± 46. 05 (μg/L) , (627.83 ±98.44,680. 17 ± 160. 18,773.50 ±80.84 pmol/L) , (1785. 02 ±321.61,2096.71 ±344.71,3430.02 ± 1538. 38 nmol/L) respectively producted by LGC,when LGC were

  2. Expression of AKT1 along with AKT2 in granulosa-lutein cells of hyperandrogenic PCOS patients.

    Science.gov (United States)

    Nekoonam, Saeid; Naji, Mohammad; Nashtaei, Maryam Shabani; Mortezaee, Keywan; Koruji, Morteza; Safdarian, Leili; Amidi, Fardin

    2017-04-01

    AKTs have a pivotal role in the granulosa-lutein cell (GC) proliferation and folliculogenesis, and there is a reciprocal feedback between AKT with androgen. Therefore, we aimed to evaluate the role of AKTs in GCs of hyperandrogenic (+HA) PCOS cases. There were three groups: control, +HA PCOS and -HA (non-hyperandrogenic) PCOS. All groups were subjected to GnRH antagonist protocol for stimulation of ovulation. Follicular fluid was aspirated from large follicles, and GCs were isolated using cell strainer method. AKT1, AKT2, AKT3, and androgen receptor (AR) mRNA expressions were analyzed with quantitative real-time PCR (qRT-PCR), and total-AKT and p-AKT (Ser(473) & Thr(308)) were investigated using western blotting. There were high levels of AKT1, AKT2, and AR mRNA expressions and high levels of p-AKT protein expression in the +HA PCOS group (p ≤ 0.05). There was a direct positive correlation between free testosterone (FT) and total testosterone (TT) with the levels of AKT1, AKT2, and p-AKT (Ser(473)), and also between FT with the levels of AR. High expressions of AKT1 and AKT2 through possible relation with androgen may cause GCs dysfunction in the +HA PCOS patients.

  3. Is Lutein a Physiologically Important Ligand for Transthyretin in Humans?

    Energy Technology Data Exchange (ETDEWEB)

    Liwei Chen

    2003-05-31

    Lutein and zeaxanthin are the only carotenoids accumulated in the macula of the human retina and are known as the macular pigments (MP). These pigments account for the yellow color of the macula and appear to play an important role in protecting against age-related macular degeneration (AMD). The uptake of lutein and zeaxanthin in human eyes is remarkably specific. It is likely that specific transport or binding proteins are involved. The objective is to determine whether transthyretin (TTR) is a transport protein in human plasma and could thus deliver lutein from the blood to the retina. In this study, they used a biosynthetic {sup 13}C-lutein tracer and gas chromatography-combustion interfaced-isotope ratio mass spectrometry (GCC-IRMS) to gain the requisite sensitivity to detect the minute amounts of lutein expected as a physiological ligand for human transthyretin. The biosynthetic {sup 13}C-labeled lutein tracer was purified from algae. Healthy women (n = 4) each ingested 1 mg of {sup 13}C-labeled lutein daily for 3 days and a blood sample was collected 24 hours after the final dose. Plasma TTR was isolated by retinol-binding protein (RBP)-sepharose affinity chromatography and extracted with chloroform. The {sup 13}C/{sup 12}C ratio in the TTR extract was measured by GCC-IRMS. There was no {sup 13}C-lutein enrichment in the pure TTR extract. This result indicated that lutein is not associated with TTR in human plasma after ingestion in physiological amounts. Some hydrophobic compounds with yellow color may bind to human TTR in the plasma. However, this association needs to be further proved by showing specificity. The study provides a new approach for carotenoid-binding protein studies using a stable isotope tracer method combined with the high precision of GCC-IRMS. The mechanism of selective transport, uptake, and accumulation of lutein in human macula remain to be determined.

  4. Is Lutein a Physiologically Important Ligand for Transthyretin in Humans?

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Liwei [Iowa State Univ., Ames, IA (United States)

    2003-01-01

    Lutein and zeaxanthin are the only carotenoids accumulated in the macula of the human retina and are known as the macular pigments (MP). These pigments account for the yellow color of the macula and appear to play an important role in protecting against age-related macular degeneration (AMD). The uptake of lutein and zeaxanthin in human eyes is remarkably specific. It is likely that specific transport or binding proteins are involved. The objective is to determine whether transthyretin (TTR) is a transport protein in human plasma and could thus deliver lutein from the blood to the retina. In this study, they used a biosynthetic 13C-lutein tracer and gas chromatography-combustion interfaced-isotope ratio mass spectrometry (GCC-IRMS) to gain the requisite sensitivity to detect the minute amounts of lutein expected as a physiological ligand for human transthyretin. The biosynthetic 13C-labeled lutein tracer was purified from algae. Healthy women (n = 4) each ingested 1 mg of 13C-labeled lutein daily for 3 days and a blood sample was collected 24 hours after the final dose. Plasma TTR was isolated by retinol-binding protein (RBP)-sepharose affinity chromatography and extracted with chloroform. The 13C/12C ratio in the TTR extract was measured by GCC-IRMS. There was no 13C-lutein enrichment in the pure TTR extract. This result indicated that lutein is not associated with TTR in human plasma after ingestion in physiological amounts. Some hydrophobic compounds with yellow color may bind to human TTR in the plasma. However, this association needs to be further proved by showing specificity. The study provides a new approach for carotenoid-binding protein studies using a stable isotope tracer method combined with the high precision of GCC-IRMS. The mechanism of selective transport, uptake, and accumulation of lutein in human macula remain to be determined.

  5. Antioxidant effect of lutein towards phospholipid hydroperoxidation in human erythrocytes.

    Science.gov (United States)

    Nakagawa, Kiyotaka; Kiko, Takehiro; Hatade, Keijiro; Sookwong, Phumon; Arai, Hiroyuki; Miyazawa, Teruo

    2009-11-01

    Peroxidised phospholipid-mediated cytotoxity is involved in the pathophysiology of many diseases; for example, phospholipid hydroperoxides (PLOOH) are abnormally increased in erythrocytes of dementia patients. Dietary carotenoids (especially xanthophylls, polar carotenoids such as lutein) have gained attention as potent inhibitors against erythrocyte phospholipid hydroperoxidation, thereby making them plausible candidates for preventing diseases (i.e. dementia). To evaluate these points, we investigated whether orally administered lutein is distributed to human erythrocytes, and inhibits erythrocyte PLOOH formation. Six healthy subjects took one capsule of food-grade lutein (9.67 mg lutein per capsule) once per d for 4 weeks. Before and during the supplementation period, carotenoids and PLOOH in erythrocytes and plasma were determined by our developed HPLC technique. The administered lutein was incorporated into human erythrocytes, and erythrocyte PLOOH level decreased after the ingestion for 2 and 4 weeks. The antioxidative effect of lutein was confirmed on erythrocyte membranes, but not in plasma. These results suggest that lutein has the potential to act as an important antioxidant molecule in erythrocytes, and it thereby may contribute to the prevention of dementia. Therefore future biological and clinical studies will be required to evaluate the efficacy as well as safety of lutein in models of dementia with a realistic prospect of its use in human therapy.

  6. Chicken granulosa cells show differential expression of epidermal growth factor (EGF) and luteinizing hormone (LH) receptor messenger RNA and differential responsiveness to EGF and LH dependent upon location of granulosa cells to the germinal disc.

    Science.gov (United States)

    Yao, H H; Bahr, J M

    2001-06-01

    Granulosa cells in the chicken follicle exhibit different phenotypes according to their location relative to the germinal disc (GD). Granulosa cells proximal to the GD (referred to as proximal granulosa cells) are more proliferative, whereas granulosa cells distal to the GD (referred to as distal granulosa cells) are more differentiated. We have shown that epidermal growth factor (EGF) derived from the GD stimulated proliferation of granulosa cells proximal to the GD, whereas extraovarian LH promoted differentiation. We tested the hypothesis that phenotypic differences of granulosa cells are the result of differential responsiveness of granulosa cells to EGF and LH. We found that both granulosa and theca layers of chicken preovulatory follicles expressed mRNA for EGF receptor (EGFr) by polymerase chain reaction (PCR) analysis. However, only the granulosa layer showed differential expression of EGFr and LH receptor (LHr) mRNA. Competitive reverse transcription-PCR revealed that proximal granulosa cells expressed more EGFr mRNA but less LHr mRNA than distal granulosa cells. In addition, proximal granulosa cells proliferated more in response to EGF than their distal counterparts. We further demonstrated that EGF decreased LHr mRNA expression by granulosa cells in a dose-dependent manner, whereas EGF and LH had no effect on EGFr mRNA expression except at one dose of LH (15 ng/ml) that stimulated EGFr mRNA expression. Our findings suggest that EGF derived from the GD influences the phenotypes of granulosa cells. Granulosa cells proximal to the GD exhibit a proliferative phenotype possibly because they are exposed to and are more responsive to GD-derived EGF. Furthermore, GD-derived EGF decreases LHr mRNA expression by proximal granulosa cells and therefore results in less differentiated granulosa cell phenotype. In contrast, granulosa cells distal to the GD are not under the influence of EGF and exhibit a more differentiated phenotype.

  7. Transcription factor p53 can regulate proliferation, apoptosis and secretory activity of luteinizing porcine ovarian granulosa cell cultured with and without ghrelin and FSH.

    Science.gov (United States)

    Sirotkin, A V; Benco, A; Tandlmajerova, A; Vasícek, D; Kotwica, J; Darlak, K; Valenzuela, F

    2008-11-01

    The aim of our in vitro experiments was to examine the role of transcription factor p53 in controlling the basic functions of ovarian cells and their response to hormonal treatments. Porcine ovarian granulosa cells, transfected and non-transfected with a gene construct encoding p53, were cultured with ghrelin and FSH (all at concentrations of 0, 1, 10, or 100 ng/ml). Accumulation of p53, of apoptosis-related (MAP3K5) and proliferation-related (cyclin B1) substances was evaluated by immunocytochemistry. The secretion of progesterone (P(4)), oxytocin (OT), prostaglandin F (PGF), and E (PGE) was measured by RIA. Transfection with the p53 gene construct promoted accumulation of this transcription factor within cells. It also stimulated the expression of a marker of apoptosis (MAP3K5). Over-expression of p53 resulted in reduced accumulation of a marker of proliferation (cyclin B1), P(4), and PGF secretion and increased OT and PGE secretion. Ghrelin, when added alone, did not affect p53 or P(4), but reduced MAP3K5 and increased PGF and PGE secretion. Over-expression of p53 reversed the effect of ghrelin on OT, caused it to be inhibitory to P(4) secretion, but did not modify its action on MAP3K5, PGF, or PGE. FSH promoted the accumulation of p53, MAP3K5, and cyclin B1; these effects were unaffected by p53 transfection. These multiple effects of the p53 gene construct on luteinizing granulosa cells, cultured with and without hormones 1) demonstrate the effects of ghrelin and FSH on porcine ovarian cell apoptosis and secretory activity, 2) confirm the involvement of p53 in promoting apoptosis and inhibiting P(4) secretion in these cells, 3) provide the first evidence that p53 suppress proliferation of ovarian cells, 4) provide the first evidence that p53 is involved in the control of ovarian peptide hormone (OT) and prostaglandin (PGF and PGE) secretion, and 5) suggest that p53 can modulate, but probably not mediate, the effects of ghrelin and FSH on the ovary.

  8. The magnitude of gonadotoxicity of chemotherapy drugs on ovarian follicles and granulosa cells varies depending upon the category of the drugs and the type of granulosa cells.

    Science.gov (United States)

    Yuksel, Aytac; Bildik, Gamze; Senbabaoglu, Filiz; Akin, Nazli; Arvas, Macit; Unal, Fehmi; Kilic, Yagmur; Karanfil, Isil; Eryılmaz, Baldan; Yilmaz, Pelin; Ozkanbaş, Can; Taskiran, Cagatay; Aksoy, Senai; Guzel, Yılmaz; Balaban, Basak; Ince, Umit; Iwase, Akira; Urman, Bulent; Oktem, Ozgur

    2015-12-01

    Do different chemotherapy drugs exert the same magnitude of cytotoxicity on dormant primordial follicles and the growing follicle fraction in the ovary in vivo and on mitotic non-luteinized and non-mitotic luteinized granulosa cells in vitro? Cyclophosphamide (alkylating agent) and cisplatin (alkylating like) impacted both primordial and pre-antral/antral follicles and both mitotic and non-mitotic granulosa cells, whereas the anti-metabolite cancer drug gemcitabine was detrimental only to pre-antral/antral follicles and mitotic non-luteinized granulosa cells. It is already known that anti-metabolite cancer drugs are less detrimental to the ovary than alkylating and alkylating like agents, such as cyclophosphamide and cisplatin. This assumption is largely based on the results of clinical reports showing lower rates of amenorrhea in women receiving anti-metabolite agent-based regimens compared with those treated with the protocols containing an alkylating drug or a platinum compound. But a quantitative comparison of gonadotoxicity with a histomorphometric proof of evidence has not been available for many chemotherapy drugs. Therefore, we combined in this study in vivo and in vitro models of human and rat origin that allows a comparative analysis of the impact of different chemotherapy agents on the ovary and granulosa cells using real-time quantitative cell indices, histomorphometry, steroidogenesis assays, and DNA damage and cell death/viability markers. We also aimed to investigate if there is a difference between mitotic and non-mitotic granulosa cells in terms of their sensitivity to the cytotoxic actions of chemotherapy drugs with different mechanisms of action. This issue has not been addressed previously. This translational research study involved in vivo analyses of ovaries in rats and in vitro analyses of granulosa cells of human and rat origin. For the in vivo assays, 54 4- to 6-week old Sprague-Dawley young female rats were randomly allocated into four

  9. Circadian Clock genes Per2 and clock regulate steroid production, cell proliferation, and luteinizing hormone receptor transcription in ovarian granulosa cells

    Energy Technology Data Exchange (ETDEWEB)

    Shimizu, Takashi, E-mail: shimizut@obihiro.ac.jp [Graduate School of Animal and Food Hygiene, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555 (Japan); Hirai, Yuko; Murayama, Chiaki; Miyamoto, Akio [Graduate School of Animal and Food Hygiene, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555 (Japan); Miyazaki, Hitoshi [Gene Research Center, University of Tsukuba, Tsukuba, Ibaraki 305-8572 (Japan); Miyazaki, Koyomi [Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology (AIST) Central 6, 1-1-1, Higashi, Tsukuba, Ibaraki 305-8566 (Japan)

    2011-08-19

    Highlights: {yields} Treatment with Per2 and Clock siRNAs decreased the number of granulosa cells and LHr expression. {yields}Per2 siRNA treatment did not stimulate the production of estradiol and expression of P450arom. {yields} Clock siRNA treatment inhibited the production of estradiol and expression of P450arom mRNA. {yields}Per2 and Clock siRNA treatment increased and unchanged, respectively, progesterone production in FSH-treated granulosa cells. {yields} The expression of StAR mRNA was increased by Per2 siRNA and unchanged by Clock siRNA. -- Abstract: Circadian Clock genes are associated with the estrous cycle in female animals. Treatment with Per2 and Clock siRNAs decreased the number of granulosa cells and LHr expression in follicle-stimulating hormone FSH-treated granulosa cells. Per2 siRNA treatment did not stimulate the production of estradiol and expression of P450arom, whereas Clock siRNA treatment inhibited the production of estradiol and expression of P450arom mRNA. Per2 and Clock siRNA treatment increased and unchanged, respectively, progesterone production in FSH-treated granulosa cells. Similarly, expression of StAR mRNA was increased by Per2 siRNA and unchanged by Clock siRNA. Our data provide a new insight that Per2 and Clock have different action on ovarian granulosa cell functions.

  10. Role of macrophage colony-stimulating factor (M-CSF) in human granulosa cells.

    Science.gov (United States)

    Xu, Song; Zhang, Zhifen; Xia, Li-Xia; Huang, Jian

    2016-12-01

    Macrophage colony-stimulating factor (M-CSF) has been proved to have a positive role in the follicular development. We investigated its effect on human granulosa cells and found that M-CSF could stimulate the production of E2. The production of FSH receptors was enhanced by M-CSF in vitro in a dose-dependent manner with or without the addition of tamoxifen (p M-CSF and its receptor (p M-CSF (p M-CSF has a role in regulating the response of granulosa cells to gonadotropins. Its function is associated with JAK/STAT-signaling pathway.

  11. Luteinizing hormone and human chorionic gonadotropin: origins of difference.

    Science.gov (United States)

    Choi, Janet; Smitz, Johan

    2014-03-01

    Luteinizing hormone (LH) and human chorionic gonadotropin (hCG) are widely recognized for their roles in ovulation and the support of early pregnancy. Aside from the timing of expression, however, the differences between LH and hCG have largely been overlooked in the clinical realm because of their similar molecular structures and shared receptor. With technologic advancements, including the development of highly purified and recombinant gonadotropins, researchers now appreciate that these hormones are not as interchangeable as once believed. Although they bind to a common receptor, emerging evidence suggests that LH and hCG have disparate effects on downstream signaling cascades. Increased understanding of the inherent differences between LH and hCG will foster more effective diagnostic and prognostic assays for use in a variety of clinical contexts and support the individualization of treatment strategies for conditions such as infertility.

  12. SIRT1 induces resistance to apoptosis in human granulosa cells by activating the ERK pathway and inhibiting NF-κB signaling with anti-inflammatory functions.

    Science.gov (United States)

    Han, Ying; Luo, Haining; Wang, Hui; Cai, Jun; Zhang, Yunshan

    2017-07-28

    SIRT1, a member of the sirtuin family, has recently emerged as a vital molecule in controlling ovarian function. The aims of the present study were to investigate SIRT1 expression and analyze SIRT1-mediated apoptosis in human granulosa cells (GCs). Human ovarian tissues were subjected to immunohistochemistry for localization of SIRT1 expression. SIRT1 knockdown in a human ovarian GC tumor line (COV434) was achieved by small interfering RNA, and the relationship between apoptosis and SIRT1 was assessed by quantitative reverse transcription polymerase chain reaction and western blotting. We further detected SIRT1 expression in human luteinized GCs. Associations among SIRT1 knockdown, SIRT1 stimulation (resveratrol) and expression of ERK1/2 and apoptotic regulatory proteins were analyzed in cell lines and luteinized GCs. Resveratrol downregulated the levels of nuclear factor (NF)-κB/p65, but this inhibitory effect was attenuated by suppressing SIRT1 activity. The NF-κB/p65 inhibitor pyrrolidine dithiocarbamate achieved similar anti-apoptosis effects. These results suggest that SIRT1 might play an anti-apoptotic role in apoptosis processes in GCs, possibly by sensing and regulating the ERK1/2 pathway, which has important clinical implications. Thus, our study provides a mechanistic link, whereby activation of SIRT1 function might help to sustain human reproduction by maintaining GCs as well as oocytes, offering a novel approach for developing a new class of therapeutic anti-inflammatory agents.

  13. Human pituitary and placental hormones control human insulin-like growth factor II secretion in human granulosa cells

    Energy Technology Data Exchange (ETDEWEB)

    Ramasharma, K.; Li, C.H.

    1987-05-01

    Human granulosa cells cultured with calf serum actively proliferated for 18-20 generations and secreted progesterone into the medium; progesterone levels appeared to decline with increase in generation number. Cells cultured under serum-free conditions secreted significant amounts of progesterone and insulin-like growth factor II (IGF-II). The progesterone secretion was enhanced by the addition of human follitropin, lutropin, and chorionic gonadotropin but not by growth hormone. These cells, when challenged to varying concentrations of human growth hormone, human chorionic somatomammotropin, human prolactin, chorionic gonadotropin, follitropin, and lutropin, secreted IGF-II into the medium as measured by specific IGF-II RIA. Among these human hormones, chorionic gonadotropin, follitropin, and lutropin were most effective in inducing IGF-II secretion from these cells. When synthetic lutropin-releasing hormone and ..cap alpha..-inhibin-92 were tested, only lutropin-releasing hormone was effective in releasing IGF-II. The results described suggest that cultured human granulosa cells can proliferate and actively secrete progesterone and IGF-II into the medium. IGF-II production in human granulosa cells was influenced by a multi-hormonal complex including human growth hormone, human chorionic somatomammotropin, and prolactin.

  14. GnRH receptors in human granulosa cells: Anatomical localization and characterization by autoradiographic study

    Energy Technology Data Exchange (ETDEWEB)

    Latouche, J.; Crumeyrolle-Arias, M.; Jordan, D.; Kopp, N.; Augendre-Ferrante, B.; Cedard, L.; Haour, F. (Institut Pasteur, Paris (France))

    1989-09-01

    The presence of receptors for GnRH in human ovary has been investigated by quantitative autoradiography. Simultaneous visualization and characterization of specific receptors on frozen sections were obtained on six pairs of human ovaries. Among them only one exhibited a large preovulatory follicle. This dominant follicle exhibited a specific and high affinity binding capacity for {sup 125}I-GnRHa exclusively localized on the granulosa cell layer. Analysis of saturation curve indicates a Kd value of 0.22 nM and Bmax of 9.6 fmol/mg protein. In contrast LH-hCG binding sites were present in all antral follicles. These data demonstrate for the first time the presence of high affinity GnRH receptors in human granulosa cells at a late stage of follicular maturation.

  15. Desmosomal plaque-associated vimentin filaments in human ovarian granulosa cell tumors of various histologic patterns.

    OpenAIRE

    Czernobilsky, B; Moll, R.; Leppien, G.; Schweikhart, G.; Franke, W W

    1987-01-01

    Proteins of intermediate-sized filaments and desmosomal plaques (desmoplakins) of four human ovarian granulosa cell tumors were studied by immunofluorescence and immunoelectron microscopy and by two-dimensional gel electrophoresis of microdissected tissue samples. All tumor cells, irrespective of their specific histologic patterns, contained both vimentin and desmoplakins. Cytokeratin-positive structures were absent or very scant in most tumor regions, but more common in trabecular, insular, ...

  16. Effects of leptin and neuropeptide Y on function of human ovarian granulosa cells in vitro

    Institute of Scientific and Technical Information of China (English)

    Song Qing; Chen Xiao-yan; Cao Zuan-sun; Mao Wen-jun

    2006-01-01

    Objective: To investigate the effects of leptin and neuropeptide Y on steroidogenesis of human ovarian granulosa cells in vitro. Methods: Human ovarian granulosa cells were isolated from follicular fluid obtained during oocyte retrieval of in vitro fertilization-embryo transfer program and cultured for 2 days with various concentration of leptin(1,10,100 ng/ml) or neuropeptide Y (1×10-6, 1×10-7, 1×10-8 mol/L) alone or both,or with the combination of human chorionic gonadotropin (hCG 0, 20 IU/L). The medium was collected for estradiol (E2) and progesterone (P) measurements. Results: (1)Whether hCG existed or not, the adding of leptin did not alter estradiol and progesterone production by human granulosa cells (P>0.05).(2) Only when the concentration of neuropeptide Y was at 1×10-7mol/L,estradiol level was lower than that in the control (P<0.05).(3) The levels of estradiol in neuropeptide Y (1×10-7mol/L) plus hCG group were significantly higher than those with neuropeptide Y alone(P<0.05). (4) In the absence of hCG, the levels of estradiol in neuropeptide Y (1×10-7mol/L)plus leptin (10 ng/ml) group were significantly higher than those with neuropeptide Y(1×10-7mol/L)alone(P<0.05).Conclusions: (1)Leptin alone produced no direct effect on secretion of E2 and P from granulosa cells in vitro.(2)Neuropeptide Y alone may inhibit the secretion of E2, but the inhibition would probably be blocked with the presentation of hCG.(3)Leptin probably blocked the inhibition of neuropeptide Y on E2 secretion, and this may indicate that there were some coordination between leptin and neuropeptid Y on the level of ovarian function.

  17. Structure of the lutein-binding domain of human StARD3 at 1.74 Å resolution and model of a complex with lutein

    Energy Technology Data Exchange (ETDEWEB)

    Horvath, Martin P., E-mail: martin.horvath@utah.edu; George, Evan W.; Tran, Quang T.; Baumgardner, Kody; Zharov, Gabe; Lee, Sarah [University of Utah, 257 S 1400 E, Salt Lake City, UT 84112 (United States); Sharifzadeh, Hassan; Shihab, Saeed; Mattinson, Ty; Li, Binxing; Bernstein, Paul S., E-mail: martin.horvath@utah.edu [Moran Eye Center, University of Utah School of Medicine, Salt Lake City, UT 84132 (United States)

    2016-07-27

    The structure of a START-domain protein known to bind lutein in the human retina is reported to an improved resolution limit. Rigid-body docking demonstrates that at least a portion of lutein must protrude from the large tunnel-like cavity characteristic of this helix-grip protein and suggests a mechanism for lutein binding specificity. A crystal structure of the lutein-binding domain of human StARD3 (StAR-related lipid-transfer protein 3; also known as MLN64) has been refined to 1.74 Å resolution. A previous structure of the same protein determined to 2.2 Å resolution highlighted homology with StARD1 and shared cholesterol-binding character. StARD3 has since been recognized as a carotenoid-binding protein in the primate retina, where its biochemical function of binding lutein with specificity appears to be well suited to recruit this photoprotective molecule. The current and previous structures correspond closely to each other (r.m.s.d. of 0.25 Å), especially in terms of the helix-grip fold constructed around a solvent-filled cavity. Regions of interest were defined with alternate conformations in the current higher-resolution structure, including Arg351 found within the cavity and Ω1, a loop of four residues found just outside the cavity entrance. Models of the complex with lutein generated by rigid-body docking indicate that one of the ionone rings must protrude outside the cavity, and this insight has implications for molecular interactions with transport proteins and enzymes that act on lutein. Interestingly, models with the ∊-ionone ring characteristic of lutein pointing towards the bottom of the cavity were associated with fewer steric clashes, suggesting that steric complementarity and ligand asymmetry may play a role in discriminating lutein from the other ocular carotenoids zeaxanthin and meso-zeaxanthin, which only have β-ionone rings.

  18. Effect of Bisphenol A on steroid production in human granulosa cells in vitro

    Institute of Scientific and Technical Information of China (English)

    Nancy Jiang; Cui Yu-gui; Zhou Wei; Liu Jin-yong; Liu Jia-yin

    2008-01-01

    Objective:To study the effects of environmental estrogen-like chemical bisphenol A(BPA)on ovarian function with the model of the cultured human granulosa cells in vitro.Methods:The granulosa cells from healthy women were collected and cultured in DMEM with 10% heat-inacti-vated fetal bovine serum.BPA,final concentration 10-7 to 10-4 mol/L,was added to the cell-culture medium and treat cells for 48 hours.The levels of estrogen and progesterone in the supernatant of the cultured cells were meas-ured by DELFIA.Total RNA was extracted from the cultured cells.The expression levels of P450 scc and P450 arom mRNA were measured by RT-PCR.Results:The cultured human granulosa cells could express high levels of P450scc and P450arom.The levels of estrogen and progesterone increased after BPA treatment,expression of P450 arom mRNA was reduced significantly at 10-5 to 10-4mol/L of BPA,but the expression of P450 scc mRNA was increased at 10-6 to 10-5 mol/L of BPA.The levels of estrogen and progesterone in the supernatant of the cultured cells were not affected significantly by BPA at the same concentrations incubated for 48 hours.Conclusion:BPA could affect steroid hormone synthesis and transformation in granulosa cells,such as the ex-pression of P450 scc and P450 atom,so it can affect ovarian function.Although we did not find a significant effect of BPA on the final estrogen and progesterone levels in this studying model,noxious effects of BPA on ovarian function may be exist in the human granulose cells.

  19. The effect of calcium phosphate nanoparticles on hormone production and apoptosis in human granulosa cells

    Directory of Open Access Journals (Sweden)

    Gao Li

    2010-04-01

    Full Text Available Abstract Objectives Although many nanomaterials are being used in academia, industry and daily life, there is little understanding about the effects of nanoparticles on the reproductive health of vertebral animals, including human beings. An experimental study was therefore performed here to explore the effect of calcium phosphate nanoparticles on both steroid hormone production and apoptosis in human ovarian granulosa cells. Methods Calcium phosphate nanoparticles uptaking was evaluated by transmission electron microscopy (TEM. The cell cycle was assessed with propidium iodide-stained cells (distribution of cells in G0/G1, S, and G2/M phases by flow cytometry. The pattern of cell death (necrosis and apoptosis was analyzed by flow cytometry with annexin V-FITC/PI staining. The expression of mRNAs encoding P450scc, P450arom and StAR were determined by RT-PCR. Progesterone and estradiol levels were measured by radioimmunoassay. Results TEM results confirmed that calcium phosphate nanoparticles could enter into granulosa cells, and distributed in the membranate compartments, including lysosome and mitochondria and intracellular vesicles. The increased percentage of cells in S phase when cultured with nanoparticles indicated that there was an arrest at the checkpoint from phase S-to-G2/M (from 6.28 +/- 1.55% to 11.18 +/- 1.73%, p Conclusion Calcium phosphate nanoparticles interfered with cell cycle of cultured human ovarian granulosa cells thus increasing cell apoptosis. This pilot study suggested that effects of nanoparticles on ovarian function should be extensively investigated.

  20. Estrogen Modulates Specific Life and Death Signals Induced by LH and hCG in Human Primary Granulosa Cells In Vitro.

    Science.gov (United States)

    Casarini, Livio; Riccetti, Laura; De Pascali, Francesco; Gilioli, Lisa; Marino, Marco; Vecchi, Eugenia; Morini, Daria; Nicoli, Alessia; La Sala, Giovanni Battista; Simoni, Manuela

    2017-04-28

    Luteinizing hormone (LH) and human chorionic gonadotropin (hCG) are glycoprotein hormones used for assisted reproduction acting on the same receptor (LHCGR) and mediating different intracellular signaling. We evaluated the pro- and anti-apoptotic effect of 100 pM LH or hCG, in the presence or in the absence of 200 pg/mL 17β-estradiol, in long-term, serum-starved human primary granulosa cells (hGLC) and a transfected granulosa cell line overexpressing LHCGR (hGL5/LHCGR). To this purpose, phospho-extracellular-regulated kinase 1/2 (pERK1/2), protein kinase B (pAKT), cAMP-responsive element binding protein (pCREB) activation and procaspase 3 cleavage were evaluated over three days by Western blotting, along with the expression of target genes by real-time PCR and cell viability by colorimetric assay. We found that LH induced predominant pERK1/2 and pAKT activation STARD1, CCND2 and anti-apoptotic XIAP gene expression, while hCG mediated more potent CREB phosphorylation, expression of CYP19A1 and procaspase 3 cleavage than LH. Cell treatment by LH is accompanied by increased (serum-starved) cell viability, while hCG decreased the number of viable cells. The hCG-specific, pro-apoptotic effect was blocked by a physiological dose of 17β-estradiol, resulting in pAKT activation, lack of procaspase 3 cleavage and increased cell viability. These results confirm that relatively high levels of steroidogenic pathway activation are linked to pro-apoptotic signals in vitro, which may be counteracted by other factors, i.e., estrogens.

  1. Exploratory Metabolomic Analyses Reveal Compounds Correlated with Lutein Concentration in Frontal Cortex, Hippocampus, and Occipital Cortex of Human Infant Brain.

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    Jacqueline C Lieblein-Boff

    Full Text Available Lutein is a dietary carotenoid well known for its role as an antioxidant in the macula, and recent reports implicate a role for lutein in cognitive function. Lutein is the dominant carotenoid in both pediatric and geriatric brain tissue. In addition, cognitive function in older adults correlated with macular and postmortem brain lutein concentrations. Furthermore, lutein was found to preferentially accumulate in the infant brain in comparison to other carotenoids that are predominant in diet. While lutein is consistently related to cognitive function, the mechanisms by which lutein may influence cognition are not clear. In an effort to identify potential mechanisms through which lutein might influence neurodevelopment, an exploratory study relating metabolite signatures and lutein was completed. Post-mortem metabolomic analyses were performed on human infant brain tissues in three regions important for learning and memory: the frontal cortex, hippocampus, and occipital cortex. Metabolomic profiles were compared to lutein concentration, and correlations were identified and reported here. A total of 1276 correlations were carried out across all brain regions. Of 427 metabolites analyzed, 257 were metabolites of known identity. Unidentified metabolite correlations (510 were excluded. In addition, moderate correlations with xenobiotic relationships (2 or those driven by single outliers (3 were excluded from further study. Lutein concentrations correlated with lipid pathway metabolites, energy pathway metabolites, brain osmolytes, amino acid neurotransmitters, and the antioxidant homocarnosine. These correlations were often brain region-specific. Revealing relationships between lutein and metabolic pathways may help identify potential candidates on which to complete further analyses and may shed light on important roles of lutein in the human brain during development.

  2. Soy promotes juvenile granulosa cell tumor development in mice and in the human granulosa cell tumor-derived COV434 cell line.

    Science.gov (United States)

    Mansouri-Attia, Nadéra; James, Rebecca; Ligon, Alysse; Li, Xiaohui; Pangas, Stephanie A

    2014-10-01

    Soy attracts attention for its health benefits, such as lowering cholesterol or preventing breast and colon cancer. Soybeans contain isoflavones, which act as phytoestrogens. Even though isoflavones have beneficial health effects, a role for isoflavones in the initiation and progression of diseases including cancer is becoming increasingly recognized. While data from rodent studies suggest that neonatal exposure to genistein (the predominant isoflavone in soy) disrupts normal reproductive function, its role in ovarian cancers, particularly granulosa cell tumors (GCT), is largely unknown. Our study aimed to define the contribution of a soy diet in GCT development using a genetically modified mouse model for juvenile GCTs (JGCT; Smad1 Smad5 conditional double knockout mice) as well as a human JGCT cell line (COV434). While dietary soy cannot initiate JGCT development in mice, we show that it has dramatic effects on GCT growth and tumor progression compared to a soy-free diet. Loss of Smad1 and Smad5 alters estrogen receptor alpha (Esr1) expression in granulosa cells, perhaps sensitizing the cells to the effects of genistein. In addition, we found that genistein modulates estrogen receptor expression in the human JGCT cell line and positively promotes cell growth in part by suppressing caspase-dependent apoptosis. Combined, our work suggests that dietary soy consumption has deleterious effects on GCT development.

  3. Observation on human ovarian theca cell and granulosa cell interaction in polycystic ovary syndrome

    Institute of Scientific and Technical Information of China (English)

    焦泽旭; 周灿权; 庄广伦; 梁晓燕

    2002-01-01

    Objective: To investigate the role of human theca cell(TC) and granulo sa cell(GC) interaction and insulin(INS) in steroidogenesis in normal ovarian cy cle and in patients with PCOS. Methods: Ovarian theca and granulosa cells from eleven normal wo men and eight PCOS patients were co-cultured on opposite side of collagen with or without INS. The concentrations of estradiol(E2), progesterone(P) and andro stenedione (A) in the culture medium were examined by ELISA method.Results: When co-cultured with GC, TC in PCOS group produced mo re A and less P than those of normal group. When co-cultured with theca cells, granulosa cells in PCOS group produced more E2 than those of normal group. Add ition of INS increased the difference significantly.Conclusions: The GC and TC interaction from the normal and PCOS ovaries is different. There is a high A and high E2 intraovary loop of PC OS leading to premature arrest of follicle growth and anovulation. Insulin may p lay an important regulatory role.

  4. Expression of betaglycan, an inhibin coreceptor, in normal human ovaries and ovarian sex cord-stromal tumors and its regulation in cultured human granulosa-luteal cells.

    Science.gov (United States)

    Liu, Jianqi; Kuulasmaa, Tiina; Kosma, Veli-Matti; Bützow, Ralf; Vänttinen, Teemu; Hydén-Granskog, Christel; Voutilainen, Raimo

    2003-10-01

    Activins and inhibins are often antagonistic in the regulation of ovarian function. TGFbeta type III receptor, betaglycan, has been identified as a coreceptor to enhance the binding of inhibins to activin type II receptor and thus to prevent the binding of activins to their receptor. In this study we characterized the expression and regulation pattern of betaglycan gene in normal ovaries and sex cord-stromal tumors and in cultured human granulosa-luteal cells from women undergoing in vitro fertilization. Expression of betaglycan mRNA was detected by RT-PCR or Northern blotting in normal ovarian granulosa, thecal, and stroma cells as well as in granulosa-luteal cells. Immunohistochemical analysis revealed positive staining for betaglycan in antral and preovulatory follicular granulosa and thecal cells and in corpora lutea of normal ovaries. Furthermore, betaglycan expression was detected in the vast majority of granulosa cell tumors, thecomas, and fibromas, with weaker staining in granulosa cell tumors compared with fibrothecomas. In cultured granulosa-luteal cells, FSH and LH treatment increased dose-dependently the accumulation of betaglycan mRNA, as did the protein kinase A activator dibutyryl cAMP and the protein kinase C inhibitor staurosporine. In contrast, the protein kinase C activator 12-O-tetradecanoyl phorbol 13-acetate had no significant effect on betaglycan mRNA levels. Treatment with prostaglandin E(2) and with its receptor EP2 subtype agonist butaprost increased betaglycan mRNA accumulation and progesterone secretion dose- and time-dependently. In summary, betaglycan gene is expressed in normal human ovarian steroidogenic cells and sex cord-stromal ovarian tumors. The accumulation of its mRNA in cultured granulosa-luteal cells is up-regulated by gonadotropins and prostaglandin E(2), probably via the protein kinase A pathway. The specific expression and regulation pattern of betaglycan gene may be related to the functional antagonism of inhibins to

  5. Follicular fluid levels of prostaglandin E2 and the effect of prostaglandin E2 on steroidogenesis in granulosa-lutein cells in women with moderate and severe endometriosis undergoing in vitro fertilization and embryo transfer

    Institute of Scientific and Technical Information of China (English)

    WANG Jing; SHEN Xin-xin; HUANG Xiang-hua; ZHAO Zhi-ming

    2012-01-01

    Background The mechanisms of endometriosis with infertility have not been fully studied.The present study aimed to assess the follicular fluid(FF)levels of prostaglandin E2(PGE2),which plays a critical role within the ovary,and to investigate the effect of PGE2 on steroidogenesis in granulosa-lutein cells(GLCs)from women with and without endometriosis.Methods Thirty-three women with laparoscopically documented endometriosis and 40 controls undergoing in vitro fertilization(IVF)were studied.We assayed the concentrations of PGE2 in FF,the production of E2 and progesterone in FF and in culture medium,and the expression of steroidogenic acute regulatory protein(StAR)and CYP19A1 in GLCs with the intervention of PGE2.Results PGE2 and progesterone concentrations were increased and displayed positive correlation in endometriotic FF.PGE2 induced the expression of StAR and the production of progesterone in GLCs from women with endometriosis,and the expression of StAR and the production of progesterone were increased in GLCs from women with endometriosis.However,there were no significant effects of PGE2 on promoting the production of E2 or the expression of CYP19A1 in GLCs.Moreover,the production of E2 and the expression of CYP19A1 in GLCs from women with endometriosis were significantly decreased compared to the controls.Conclusions PGE2 concentrations are increased in endometriotic FF,along with concomitant increases in progesterone and StAR.In contrast,the E2 and CYP19A1 are decreased in GLCs,which may delay the development of the follicles and cause an imbalance in the follicular steroid hormone levels.These changes may have close relationship with endometriosis-associated infertility.

  6. TLR4 activates NF-{kappa}B in human ovarian granulosa tumor cells

    Energy Technology Data Exchange (ETDEWEB)

    Woods, Dori C., E-mail: dwoods2@partners.org [Vincent Center for Reproductive Biology, Vincent Obstetrics and Gynecology Service, Massachusetts General Hospital/Harvard Medical School, Boston, MA 02114 (United States); White, Yvonne A.R. [Vincent Center for Reproductive Biology, Vincent Obstetrics and Gynecology Service, Massachusetts General Hospital/Harvard Medical School, Boston, MA 02114 (United States); Dau, Caroline [University of California, San Francisco, School of Dentistry, San Francisco, CA 94143 (United States); Johnson, A.L. [Center for Reproductive Biology and Health, The Pennsylvania State University, University Park, PA 16802 (United States)

    2011-06-17

    Highlights: {yields} TLR4 is expressed in human ovarian granulosa tumor cells. {yields} Acting through TLR4, LPS and HSP60 induce a NF{kappa}B signaling cascade in human ovarian granulosa tumor cells. {yields} NF{kappa}B activation or inhibition did not alter chemosensitivity to TRAIL or cisplatin. -- Abstract: Previous studies have demonstrated expression of Toll-like receptors (TLRs) in the surface epithelium of normal ovaries (OSE) and in epithelial ovarian tumors. Most notably, OSE-derived cancers express TLR4, which activates the nuclear factor-kappa B (NF-{kappa}B) signaling cascade as a mediator of inflammatory response. Currently, there is considerable interest in elucidating the role of TLR-mediated signaling in cancers. Nevertheless, the expression of TLRs in granulosa cell tumors (GCTs) of the ovary, and the extent to which GCT expression of TLRs may influence cell-signaling pathways and/or modulate the efficacy of chemotherapeutics, has yet to be determined. In the present study, human GCT lines (COV434 and KGN) were utilized to evaluate expression of functional TLR4. TLR4 is expressed in GCT cell lines and ligation of TLR4 with bacterial lipopolysaccharide (LPS) led to I{kappa}B degradation and activation of NF-{kappa}B. NF-{kappa}B activation was confirmed by nuclear localization of NF-{kappa}B p65 following treatment with LPS and the naturally occurring ligand, HSP60. Notably, immunoneutralization of TLR4 blocked nuclear localization, and inhibition of NF-{kappa}B signaling attenuated LPS-induced TNF{alpha} plus increased doubling time in both cell lines. Contradictory to reports using human OSE cell lines, inhibition of NF-{kappa}B signaling failed to sensitize GCT lines to TRAIL or cisplatin. In summary, findings herein are the first to demonstrate a functional TLR-signaling pathway specifically in GCTs, and indicate that in contrast to OSE-derived cancers, inhibition of NF-{kappa}B does not sensitize GCTs to TRAIL or cisplatin.

  7. Lutein and Brain Function

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    John W. Erdman

    2015-10-01

    Full Text Available Lutein is one of the most prevalent carotenoids in nature and in the human diet. Together with zeaxanthin, it is highly concentrated as macular pigment in the foveal retina of primates, attenuating blue light exposure, providing protection from photo-oxidation and enhancing visual performance. Recently, interest in lutein has expanded beyond the retina to its possible contributions to brain development and function. Only primates accumulate lutein within the brain, but little is known about its distribution or physiological role. Our team has begun to utilize the rhesus macaque (Macaca mulatta model to study the uptake and bio-localization of lutein in the brain. Our overall goal has been to assess the association of lutein localization with brain function. In this review, we will first cover the evolution of the non-human primate model for lutein and brain studies, discuss prior association studies of lutein with retina and brain function, and review approaches that can be used to localize brain lutein. We also describe our approach to the biosynthesis of 13C-lutein, which will allow investigation of lutein flux, localization, metabolism and pharmacokinetics. Lastly, we describe potential future research opportunities.

  8. In human granulosa cells from small antral follicles, androgen receptor mRNA and androgen levels in follicular fluid correlate with FSH receptor mRNA

    DEFF Research Database (Denmark)

    Nielsen, M. E.; Rasmussen, I. A.; Kristensen, Stine Gry

    2011-01-01

    concentrations of AMH, inhibin-B, progesterone and estradiol. Androgen Receptor mRNA expression in granulosa cells, and the FF content of androgens, both showed a highly significant positive association with to the expression of FSHR mRNA in granulosa cells. AR mRNA expression also correlated significantly...... with the expression of AMHR2, but did not correlate with any of the hormones in the FF. These data demonstrate an intimate association between AR expression in immature granulosa cells, and the expression of FSHR in normal small human antral follicles and between the FF levels of androgen and FSHR expression...

  9. Leptin interferes with 3',5'-Cyclic Adenosine Monophosphate (cAMP signaling to inhibit steroidogenesis in human granulosa cells

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    HoYuen Basil

    2009-10-01

    Full Text Available Abstract Background Obesity has been linked to an increased risk of female infertility. Leptin, an adipocytokine which is elevated during obesity, may influence gonadal function through modulating steroidogenesis in granulosa cells. Methods The effect of leptin on progesterone production in simian virus 40 immortalized granulosa (SVOG cells was examined by Enzyme linked immunosorbent assay (ELISA. The effect of leptin on the expression of the steroidogenic enzymes (StAR, P450scc, 3betaHSD in SVOG cells was examined by real-time PCR and Western blotting. The mRNA expression of leptin receptor isoforms in SVOG cells were examined by using PCR. SVOG cells were co-treated with leptin and specific pharmacological inhibitors to identify the signaling pathways involved in leptin-reduced progesterone production. Silencing RNA against leptin receptor was used to determine that the inhibition of leptin on cAMP-induced steroidogenesis acts in a leptin receptor-dependent manner. Results and Conclusion In the present study, we investigated the cellular mechanisms underlying leptin-regulated steroidogenesis in human granulosa cells. We show that leptin inhibits 8-bromo cAMP-stimulated progesterone production in a concentration-dependent manner. Furthermore, we show that leptin inhibits expression of the cAMP-stimulated steroidogenic acute regulatory (StAR protein, the rate limiting de novo protein in progesterone synthesis. Leptin induces the activation of ERK1/2, p38 and JNK but only the ERK1/2 (PD98059 and p38 (SB203580 inhibitors attenuate the leptin-induced inhibition of cAMP-stimulated StAR protein expression and progesterone production. These data suggest that the leptin-induced MAPK signal transduction pathway interferes with cAMP/PKA-stimulated steroidogenesis in human granulosa cells. Moreover, siRNA mediated knock-down of the endogenous leptin receptor attenuates the effect of leptin on cAMP-induced StAR protein expression and progesterone

  10. Differences in gene expression of granulosa cells from women undergoing controlled ovarian hyperstimulation with either recombinant follicle-stimulating hormone or highly purified human menopausal gonadotropin

    DEFF Research Database (Denmark)

    Grøndahl, Marie Louise; Borup, Rehannah; Lee, Young Bae

    2009-01-01

    OBJECTIVE: To investigate the differences in the gene expression profile of granulosa cells from preovulatory follicles after controlled ovarian hyperstimulation (COH) with recombinant follicle-stimulating hormone (FSH) or urinary human menopausal gonadotropin (hMG) FSH. DESIGN: Prospective rando...... be important for periovulatory events, which suggests that the preparation used for COH is important for granulosa cell function and may influence the developmental competence of the oocyte and the function of corpus luteum....

  11. Relationship between Concentrations of Lutein and StARD3 among Pediatric and Geriatric Human Brain Tissue.

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    Jirayu Tanprasertsuk

    Full Text Available Lutein, a dietary carotenoid, selectively accumulates in human retina and brain. While many epidemiological studies show evidence of a relationship between lutein status and cognitive health, lutein's selective uptake in human brain tissue and its potential function in early neural development and cognitive health have been poorly evaluated at a molecular level. The objective of this study was to evaluate the cross-sectional relationship between concentrations of brain lutein and StARD3 (identified as its binding protein in retinal tissue among three age groups: infants (1-4 months, n = 10, older adults (55-86 years, n = 8, and centenarians (98-105 years, n = 10. Brain lutein concentrations were analyzed by high-performance liquid chromatography and StARD3 levels were analyzed by Western Blot analysis. The strong relationship in infant brains (r = 0.75, P 0.05, seven of whom had mild cognitive impairment (MCI or dementia. These exploratory findings suggest an age-related decrease or abnormality of StARD3 activity in human brain. Given that StARD3 is also involved in cholesterol transportation, a process that is aberrant in neurodegenerative diseases, the potential protective function of lutein against these diseases remains to be explored.

  12. Growth differentiation factor 8 suppresses cell proliferation by up-regulating CTGF expression in human granulosa cells.

    Science.gov (United States)

    Chang, Hsun-Ming; Pan, Hui-Hui; Cheng, Jung-Chien; Zhu, Yi-Min; Leung, Peter C K

    2016-02-15

    Connective tissue growth factor (CTGF) is a matricellular protein that plays a critical role in the development of ovarian follicles. Growth differentiation factor 8 (GDF8) is mainly, but not exclusively, expressed in the mammalian musculoskeletal system and is a potent negative regulator of skeletal muscle growth. The aim of this study was to investigate the effects of GDF8 and CTGF on the regulation of cell proliferation in human granulosa cells and to examine its underlying molecular determinants. Using dual inhibition approaches (inhibitors and small interfering RNAs), we have demonstrated that GDF8 induces the up-regulation of CTGF expression through the activin receptor-like kinase (ALK)4/5-mediated SMAD2/3-dependent signaling pathways. In addition, the increase in CTGF expression contributes to the GDF8-induced suppressive effect on granulosa cell proliferation. Our findings suggest that GDF8 and CTGF may play critical roles in the regulation of proliferative events in human granulosa cells. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  13. Progesterone-receptor antagonists and statins decrease de novo cholesterol synthesis and increase apoptosis in rat and human periovulatory granulosa cells in vitro.

    Science.gov (United States)

    Rung, Emilia; Friberg, P Anders; Shao, Ruijin; Larsson, D G Joakim; Nielsen, Eva Ch; Svensson, Per-Arne; Carlsson, Björn; Carlsson, Lena M S; Billig, Håkan

    2005-03-01

    Progesterone-receptor (PR) stimulation promotes survival in rat and human periovulatory granulosa cells. To investigate the mechanisms involved, periovulatory rat granulosa cells were incubated in vitro with or without the PR-antagonist Org 31710. Org 31710 caused the expected increase in apoptosis, and expression profiling using cDNA microarray analysis revealed regulation of several groups of genes with functional and/or metabolic connections. This regulation included decreased expression of genes involved in follicular rupture, increased stress responses, decreased angiogenesis, and decreased cholesterol synthesis. A decreased cholesterol synthesis was verified in experiments with both rat and human periovulatory granulosa cells treated with the PR-antagonists Org 31710 or RU 486 by measuring incorporation of [14C]acetate into cholesterol, cholesterol ester, and progesterone. Correspondingly, specific inhibition of cholesterol synthesis in periovulatory rat granulosa cells using 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors (lovastatin, mevastatin, or simvastatin) increased apoptosis, measured as DNA fragmentation and caspase-3/7 activity. The increase in apoptosis caused by simvastatin was reversed by addition of the cholesterol synthesis-intermediary mevalonic acid. These results show that PR antagonists reduce cholesterol synthesis in periovulatory granulosa cells and that cholesterol synthesis is important for granulosa cell survival.

  14. Testosterona e gonadotrofina coriônica humana estimulam a esteroidogênese em células da granulosa de folículo pré-ovulatório de égua? Do testosterone and human chorionic gonadotropin stimulate steroidogenesis in granulosa cells of preovulatory follicle in mare?

    Directory of Open Access Journals (Sweden)

    M.C. Caldas-Bussiere

    2005-02-01

    Full Text Available Avaliou-se o papel da gonadotrofina coriônica humana (hCG e da testosterona na produção de progesterona (P4 e 17beta -estradiol (E2 pelas células da granulosa cultivadas in vitro de folículo antral de égua. Os tratamentos usados foram: 1- controle (nenhum hormônio adicionado, 2- 1UI hCG (0,3mig/ml e 3- 10UI hCG (3,0mig/ml. O tratamento com hCG foi realizado na presença ou não de testosterona (144ng/ml. O meio foi coletado e substituído com 0,25, 3, 6, 12, 24 e 144h de cultivo. As concentrações de P4 e E2 foram mensuradas por radioimunoensaio. Não se observou diferença entre os tratamentos 1 e 3 quanto à produção de P4 e E2; o tratamento 1 resultou em aumento da concentração de progesterona após 24h de cultura (PThe role of the human chorionic gonadotropin (hCG and testosterone was evaluated in the progesterone (P4 and estradiol-17beta (E2 production by granulosa cells of antral follicles from mare cultivated in vitro. The treatment (groups with gonadotropin consisted of: 1- control (no added hormone; 2- 1 IU hCG (0.3mg/ml and 3- 10 IU hCG (3.0mg/ml. The treatment with hCG was carried out in the presence or not of testosterone (144ng/ml. The culture medium was collected and replaced at 0.25, 3, 6, 12, 24 and 144h of culture. The concentrations of P4 and E2 were measured by radioimunoassay. Analyses of variance were used for P4 and E2, and mean of the factors were compared by the Tukey test at 5% of probability. No difference was observed between 1 and 2 groups. Treatment with 1 IU of hCG increased progesterone concentration after 24h of culture (P<0.01, only in the presence of testosterone. The concentration of estradiol increased in the presence of testosterone, reaching maximum concentration with 6h of culture (P<0.01, and reduced gradually until the observed concentration at 0.25h of culture. The addition of hCG had no effect in the synthesis of this steroid. The testosterone modulates the action of the luteinizing hormone

  15. Differential gene expression in human granulosa cells from recombinant FSH versus human menopausal gonadotropin ovarian stimulation protocols

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    Bietz Mandi G

    2010-03-01

    Full Text Available Abstract Background The study was designed to test the hypothesis that granulosa cell (GC gene expression response differs between recombinant FSH and human menopausal gonadotropin (hMG stimulation regimens. Methods Females Results After exclusions, 1736 genes exhibited differential expression between groups. Over 400 were categorized as signal transduction genes, ~180 as transcriptional regulators, and ~175 as enzymes/metabolic genes. Expression of selected genes was confirmed by RT-PCR. Differentially expressed genes included A kinase anchor protein 11 (AKAP11, bone morphogenetic protein receptor II (BMPR2, epidermal growth factor (EGF, insulin-like growth factor binding protein (IGFBP-4, IGFBP-5, and hypoxia-inducible factor (HIF-1 alpha. Conclusions Results suggest that major differences exist in the mechanism by which pure FSH alone versus FSH/LH regulate gene expression in preovulatory GC that could impact oocyte maturity and developmental competence.

  16. Identification and characterization of Ca2+-activated K+ channels in granulosa cells of the human ovary

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    Berg Ulrike

    2009-04-01

    Full Text Available Abstract Background Granulosa cells (GCs represent a major endocrine compartment of the ovary producing sex steroid hormones. Recently, we identified in human GCs a Ca2+-activated K+ channel (KCa of big conductance (BKCa, which is involved in steroidogenesis. This channel is activated by intraovarian signalling molecules (e.g. acetylcholine via raised intracellular Ca2+ levels. In this study, we aimed at characterizing 1. expression and functions of KCa channels (including BKCa beta-subunits, and 2. biophysical properties of BKCa channels. Methods GCs were obtained from in vitro-fertilization patients and cultured. Expression of mRNA was determined by standard RT-PCR and protein expression in human ovarian slices was detected by immunohistochemistry. Progesterone production was measured in cell culture supernatants using ELISAs. Single channels were recorded in the inside-out configuration of the patch-clamp technique. Results We identified two KCa types in human GCs, the intermediate- (IK and the small-conductance KCa (SK. Their functionality was concluded from attenuation of human chorionic gonadotropin-stimulated progesterone production by KCa blockers (TRAM-34, apamin. Functional IK channels were also demonstrated by electrophysiological recording of single KCa channels with distinctive features. Both, IK and BKCa channels were found to be simultaneously active in individual GCs. In agreement with functional data, we identified mRNAs encoding IK, SK1, SK2 and SK3 in human GCs and proteins of IK and SK2 in corresponding human ovarian cells. Molecular characterization of the BKCa channel revealed the presence of mRNAs encoding several BKCa beta-subunits (beta2, beta3, beta4 in human GCs. The multitude of beta-subunits detected might contribute to variations in Ca2+ dependence of individual BKCa channels which we observed in electrophysiological recordings. Conclusion Functional and molecular studies indicate the presence of active IK and SK

  17. Participation of Mitogen-activated Protein Kinase in Luteinizing Hormone-induced Differential Regulation of Steroidogenesis and Steroidogenic Gene Expression in Mural and Cumulus Granulosa Cells of Mouse Preovulatory Follicles

    DEFF Research Database (Denmark)

    Su, You-Qiang; Nyegaard, Mette; Overgaard, Michael Toft;

    2006-01-01

    was to investigate whether these processes that commonly occur in mural granulosa cells (MGCs) also occur in cumulus cells, and whether they are mediated by the mitogen-activated protein kinase (MAPK), specifically MAPK3/1 (also commonly known as extracellular signal-regulated kinase 1&2, ERK1/2). The standard...

  18. Expression of neurokinin B/NK3 receptor and kisspeptin/KISS1 receptor in human granulosa cells.

    Science.gov (United States)

    García-Ortega, J; Pinto, F M; Fernández-Sánchez, M; Prados, N; Cejudo-Román, A; Almeida, T A; Hernández, M; Romero, M; Tena-Sempere, M; Candenas, L

    2014-12-01

    Are neurokinin B (NKB), NK3 receptor (NK3R), kisspeptin (KISS1) and kisspeptin receptor (KISS1R) expressed in human ovarian granulosa cells? The NKB/NK3R and kisspeptin/KISS1R systems are co-expressed and functionally active in ovarian granulosa cells. The NKB/NK3R and KISS1/KISS1R systems are essential for reproduction. In addition to their well-recognized role in hypothalamic neurons, these peptide systems may contribute to the control of fertility by acting directly on the gonads, but such a direct gonadal role remains largely unknown. This study analyzed matched mural granulosa cells (MGCs) and cumulus cells (CCs) collected from preovulatory follicles of oocyte donors at the time of oocyte retrieval. The samples were provided by 56 oocyte donor women undergoing ovarian stimulation treatment. Follicular fluid samples containing MGCs and cumulus-oocyte complexes were collected after transvaginal ultrasound-guided oocyte retrieval. RT-PCR, quantitative real-time PCR, immunocytochemistry and western blot were used to investigate the pattern of expression of the NKB/NK3R and KISS/KISS1R systems in MGCs and CCs. Intracellular free Ca(2+) levels, [Ca(2+)]i, in MGCs after exposure to NKB or KISS1, in the presence or not of tachykinin receptor antagonists, were also measured. NKB/NK3R and KISS1/KISS1R systems were expressed, at the mRNA and protein levels, in MGCs and CCs, with significantly higher expression in CCs. Kisspeptin increased the [Ca(2+)]i in the cytosol of human MGCs while exposure to NKB failed to induce any change in [Ca(2+)]i. However, the [Ca(2+)]i response to kisspeptin was reduced in the presence of NKB. The inhibitory effect of NKB was only partially mimicked by the NK3R agonist, senktide and marginally suppressed by the NK3R-selective antagonist SB 222200. Yet, a cocktail of antagonists selective for the NK1, NK2 and NK3 receptors blocked the effect of NKB. The granulosa and cumulus cells were obtained from oocyte donors undergoing ovarian

  19. Amphiregulin mediates hCG-induced StAR expression and progesterone production in human granulosa cells

    Science.gov (United States)

    Fang, Lanlan; Yu, Yiping; Zhang, Ruizhe; He, Jingyan; Sun, Ying-Pu

    2016-01-01

    Progesterone plays critical roles in maintaining a successful pregnancy at the early embryonic stage. Human chorionic gonadotropin (hCG) rapidly induces amphiregulin (AREG) expression. However, it remains unknown whether AREG mediates hCG-induced progesterone production. Thus, the objective of this study was to investigate the role of AREG in hCG-induced progesterone production and the underlying molecular mechanism in human granulosa cells; primary cells were used as the experimental model. We demonstrated that the inhibition of EGFR and the knockdown of AREG abolished hCG-induced steroidogenic acute regulatory protein (StAR) expression and progesterone production. Importantly, follicular fluid AREG levels were positively correlated with progesterone levels in the follicular fluid and serum. Treatment with AREG increased StAR expression and progesterone production, and these stimulatory effects were abolished by EGFR inhibition. Moreover, activation of ERK1/2, but not PI3K/Akt, signaling was required for the AREG-induced up-regulation of StAR expression and progesterone production. Our results demonstrate that AREG mediates hCG-induced StAR expression and progesterone production in human granulosa cells, providing novel evidence for the role of AREG in the regulation of steroidogenesis. PMID:27113901

  20. CD9 Expression by Human Granulosa Cells and Platelets as a Predictor of Fertilization Success during IVF

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    Carolyn R. Jaslow

    2010-01-01

    Full Text Available Objective. To determine whether CD9 expression on human granulosa cells (GCs and platelets could predict the success of conventional fertilization of human oocytes during in vitro fertilization (IVF. Methods. Thirty women undergoing IVF for nonmale factor infertility participated. Platelets from venous blood and GCs separated from retrieved oocytes were prepared for immunofluorescence. Flow cytometry quantified the percent of GCs expressing CD9, and CD9 surface density on GCs and platelets. Fertilization rate was determined for the total number of oocytes, and the number of mature oocytes per patient. Correlations tested for significant relationships (P<.05 between fertilization rates and CD9 expression. Results. CD9 surface density on human GCs is inversely correlated with fertilization rate of oocytes (P=.04, but the relationship was weak. Conclusion. More studies are needed to determine if CD9 expression on GCs would be useful for predicting conventional fertilization success during IVF.

  1. Involvement of ERK1/2 signaling pathway in atrazine action on FSH-stimulated LHR and CYP19A1 expression in rat granulosa cells

    Energy Technology Data Exchange (ETDEWEB)

    Fa, Svetlana; Pogrmic-Majkic, Kristina; Samardzija, Dragana; Glisic, Branka; Kaisarevic, Sonja; Kovacevic, Radmila; Andric, Nebojsa, E-mail: nebojsa.andric@dbe.uns.ac.rs

    2013-07-01

    Worldwide used herbicide atrazine is linked to reproductive dysfunction in females. In this study, we investigated the effects and the mechanism of atrazine action in the ovary using a primary culture of immature granulosa cells. In granulosa cells, follicle-stimulating hormone (FSH) activates both cyclic adenosine monophosphate (cAMP) and extracellular-regulated kinase 1/2 (ERK1/2) cascades, with cAMP pathway being more important for luteinizing hormone receptor (LHR) and aromatase (CYP19A1) mRNA expression. We report that 48 h after atrazine exposure the FSH-stimulated LHR and CYP19A1 mRNA expression and estradiol synthesis were decreased, with LHR mRNA being more sensitive to atrazine than CYP19A1 mRNA. Inadequate acquisition of LHR in the FSH-stimulated and atrazine-exposed granulosa cells renders human chorionic gonadotropin (hCG) ineffective to stimulate amphiregulin (Areg), epiregulin (Ereg), and progesterone receptor (Pgr) mRNA expression, suggesting anti-ovulatory effect of atrazine. To dissect the signaling cascade involved in atrazine action in granulosa cells, we used U0126, a pharmacological inhibitor of ERK1/2. U0126 prevents atrazine-induced decrease in LHR and CYP19A1 mRNA levels and estradiol production in the FSH-stimulated granulosa cells. ERK1/2 inactivation restores the ability of hCG to induce expression of the ovulatory genes in atrazine-exposed granulosa cells. Cell-based ELISA assay revealed that atrazine does not change the FSH-stimulated ERK1/2 phosphorylation in granulosa cells. The results from this study reveal that atrazine does not affect but requires ERK1/2 phosphorylation to cause decrease in the FSH-induced LHR and CYP19A1 mRNA levels and estradiol production in immature granulosa cells, thus compromising ovulation and female fertility. - Highlights: • Atrazine inhibits estradiol production in FSH-stimulated granulosa cells. • Atrazine inhibits LHR and Cyp19a1 mRNA expression in FSH-stimulated granulosa cells. • Atrazine

  2. Specific genes are selectively expressed between cumulus and granulosa cells from individual human pre-ovulatory follicles

    DEFF Research Database (Denmark)

    Grøndahl, M L; Andersen, C Yding; Bogstad, J;

    2012-01-01

    During folliculogenesis the granulosa cells differentiate into two cell types: Cumulus cells (CC) and mural granulosa cells (MGC). The objective of the study was to generate and compare the transcriptomes of MGC and CC from the pre-ovulatory follicle to characterize the detailed profile of the two...

  3. Radioimmunoassay of bovine, ovine and porcine luteinizing hormone with a monoclonal antibody and a human tracer

    Energy Technology Data Exchange (ETDEWEB)

    Fosberg, M.; Tagle, R.; Madej, A.; Molina, J.R.; Carlsson, M.-A.

    1993-01-01

    A radioimmunoassay for bovine (bLH), ovine (oLH) and porcine (pLH) luteinizing hormone was developed using a human [sup 125]ILH tracer from a commercial kit and a monoclonal antibody (518B7) specific for LH but with low species specificity. Standard curves demonstrated similar binding kinetics when bLH, oLH and pLH were incubated with tracer and antibody for 2 h at room temperature. A 30-min delay in the addition of the tracer gave sufficient sensitivity when analysing pLH. Separation of antibody-bound LH from free hormone was achieved by using second antibody-coated micro Sepharose beads. The assay was validated and the performance compared with that of an RIA currently in use for determination of bLH (coefficient of correlation: 0.99 and 0.98). Regardless of the standards used, intra-assay coefficients of variation were <10% for LH concentrations exceeding 1 [mu]g/L. The inter-assay coefficients of variation were <15%. The assay was used for clinical evaluation demonstrating the pre-ovulatory LH surge in two cyclic cows, LH pulsatility in an oophorectomized ewe and LH response to GnRH injection in a boar. (au) (7 refs.).

  4. Growth Hormone Releasing Peptide-2 Attenuation of Protein Kinase C-Induced Inflammation in Human Ovarian Granulosa Cells

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    Yi-Ning Chao

    2016-08-01

    Full Text Available Cyclooxygenase-2 (COX-2 and interleukin-8 (IL-8 are two important inflammatory mediators in ovulation. Ghrelin may modulate inflammatory signaling via growth hormone secretagogue receptors. We investigated the role of ghrelin in KGN human ovarian granulosa cells using protein kinase C (PKC activator phorbol 12, 13-didecanoate (PDD and synthetic ghrelin analog growth hormone releasing peptide-2 (GHRP-2. GHRP-2 attenuated PDD-induced expression of protein and mRNA, the promoter activity of COX-2 and IL-8 genes, and the secretion of prostaglandin E2 (PGE2 and IL-8. GHRP-2 promoted the degradation of PDD-induced COX-2 and IL-8 proteins with the involvement of proteasomal and lysosomal pathways. PDD-mediated COX-2 production acts via the p38, c-Jun N-terminal kinase (JNK, extracellular signal-regulated kinase (ERK and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB pathways; PDD-mediated IL-8 production acts via the p38, JNK and ERK pathways. GHRP-2 reduced the PDD-induced phosphorylation of p38 and JNK and activator protein 1 (AP-1 reporter activation and PDD-induced NF-κB nuclear translocation and reporter activation. The inhibitors of mitogen-activated protein kinase phosphatase-1 (MKP-1 and protein phosphatase 2 (PP2A reduced the inhibitory effect of GHRP-2 on PDD-induced COX-2 and IL-8 expression. Our findings demonstrate an anti-inflammatory role for ghrelin (GHRP-2 in PKC-mediated inflammation of granulosa cells, at least in part, due to its inhibitory effect on PKC-induced activation of p38, JNK and NF-κB, possibly by targeting to MKP-1 and PP2A.

  5. Effects of mayonnaise on postprandial serum lutein/zeaxanthin and beta-carotene concentrations in humans.

    Science.gov (United States)

    Takeda, Sayaka; Masuda, Yasunobu; Usuda, Mika; Marushima, Ranko; Ueji, Toshiyuki; Hasegawa, Mineo; Maruyama, Chizuko

    2009-12-01

    To clarify the effects of different physical forms of oil on postprandial serum lutein/zeaxanthin and beta-carotene concentrations, we performed a vegetable meal loading test. Eighteen healthy subjects participated in the test, which consisted of broccoli as a control (CON) meal, broccoli with oil (OIL), and broccoli with mayonnaise (MS), consumed in random order. After collection of fasting blood samples, subjects consumed one of the three test meals. Fasting and postprandial changes in serum carotenoids were assessed 2, 4, and 6 h after ingestion of each test meal. Serum lutein/zeaxanthin and beta-carotene concentrations were measured. Although no significant change was noted after the CON meal, the serum lutein/zeaxanthin concentration was higher at 4 h after consumption of the OIL meal, and at 2, 4 and 6 h after consumption of the MS meal, as compared with the fasting state. Serum beta-carotene concentrations did not change after ingestion of either the CON or the OIL meal but were elevated 2, 4, and 6 h after MS ingestion as compared with the fasting state. The incremental areas under the curves (IAUCs) of serum lutein/zeaxanthin and beta-carotene concentrations were higher after the MS meal than after the CON meal. IAUCs after the OIL meal exhibited no statistically significant differences from the CON and MS meals. We suggest that mayonnaise contributes to increase serum lutein/zeaxanthin and beta-carotene concentrations when consumed with vegetables rich in these carotenoids.

  6. MicroRNA expression and regulation in human ovarian carcinoma cells by luteinizing hormone.

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    Juan Cui

    Full Text Available BACKGROUND: MicroRNAs have been widely-studied with regard to their aberrant expression and high correlation with tumorigenesis and progression in various solid tumors. With the major goal of assessing gonadotropin (luteinizing hormone, LH contributions to LH receptor (LHR-positive ovarian cancer cells, we have conducted a genome-wide transcriptomic analysis on human epithelial ovarian cancer cells to identify the microRNA-associated cellular response to LH-mediated activation of LHR. METHODS: Human ovarian cancer cells (SKOV3 were chosen as negative control (LHR- and stably transfected to express functional LHR (LHR+, followed by incubation with LH (0-20 h. At different times of LH-mediated activation of LHR the cancer cells were analyzed by a high-density Ovarian Cancer Disease-Specific-Array (DSA, ALMAC™, which profiled ∼ 100,000 transcripts with ∼ 400 non-coding microRNAs. FINDINGS: In total, 65 microRNAs were identified to exhibit differential expression in either LHR expressing SKOV3 cells or LH-treated cells, a few of which have been found in the genomic fragile regions that are associated with abnormal deletion or amplification in cancer, such as miR-21, miR-101-1, miR-210 and miR-301a. By incorporating the dramatic expression changes observed in mRNAs, strong microRNA/mRNA regulatory pairs were predicted through statistical analyses coupled with collective computational prediction. The role of each microRNA was then determined through a functional analysis based on the highly-confident microRNA/mRNA pairs. CONCLUSION: The overall impact on the transcriptome-level expression indicates that LH may regulate apoptosis and cell growth of LHR+ SKOV3 cells, particularly by reducing cancer cell proliferation, with some microRNAs involved in regulatory roles.

  7. Resveratrol promotes expression of SIRT1 and StAR in rat ovarian granulosa cells: an implicative role of SIRT1 in the ovary

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    Morita Yoshihiro

    2012-02-01

    Full Text Available Abstract Background Resveratrol is a natural polyphenolic compound known for its beneficial effects on energy homeostasis, and it also has multiple properties, including anti-oxidant, anti-inflammatory, and anti-tumor activities. Recently, silent information regulator genes (Sirtuins have been identified as targets of resveratrol. Sirtuin 1 (SIRT1, originally found as an NAD+-dependent histone deacetylase, is a principal modulator of pathways downstream of calorie restriction, and the activation of SIRT1 ameliorates glucose homeostasis and insulin sensitivity. To date, the presence and physiological role of SIRT1 in the ovary are not known. Here we found that SIRT1 was localized in granulosa cells of the human ovary. Methods The physiological roles of resveratrol and SIRT1 in the ovary were analyzed. Immunohistochemistry was performed to localize the SIRT1 expression. SIRT1 protein expression of cultured cells and luteinized human granulosa cells was investigated by Western blot. Rat granulosa cells were obtained from diethylstilbestrol treated rats. The cells were treated with increasing doses of resveratrol, and subsequently harvested to determine mRNA levels and protein levels. Cell viability was tested by MTS assay. Cellular apoptosis was analyzed by caspase 3/7 activity test and Hoechst 33342 staining. Results SIRT1 protein was expressed in the human ovarian tissues and human luteinized granulosa cells. We demonstrated that resveratrol exhibited a potent concentration-dependent inhibition of rat granulosa cells viability. However, resveratrol-induced inhibition of rat granulosa cells viability is independent of apoptosis signal. Resveratrol increased mRNA levels of SIRT1, LH receptor, StAR, and P450 aromatase, while mRNA levels of FSH receptor remained unchanged. Western blot analysis was consistent with the results of quantitative real-time RT-PCR assay. In addition, progesterone secretion was induced by the treatment of resveratrol

  8. Lutein and brain function

    Science.gov (United States)

    Lutein is one of the most prevalent carotenoids in nature and in the human diet. Together with zeaxanthin, it is highly concentrated in macular pigment in the foveal retina of primates, attenuating blue light exposure, providing protection from photo-oxidation and enhancing visual performance. Rece...

  9. Effect of gonadotr opin-releasing hormone agonist on steroidogenesis of cultured human luteinized g ranulosa cells in vitro.%GnRH-a体外对人卵巢黄素化颗粒细胞雌二醇和孕酮分泌量的影响

    Institute of Scientific and Technical Information of China (English)

    陈丹青; 黄荷凤; 朱依敏

    2001-01-01

    目的:观察促性 腺素释放激素 激动剂(GnRH-a)在体外对人卵巢黄素化颗粒细胞雌二醇(E2)和孕酮(P)分泌量的影响。方法:培养人卵巢黄素化颗粒细胞,分别用终浓度为1.0、10.0、100.0ng/ml的G nRH -a刺激,同时设对照组,培养时间为2、4、6d。用放射免疫法检测黄素化颗粒细胞E2和P 的分 泌量。结果:培养2d,GnRH-a中、高浓度组E2和P分泌量分别为(122±3 7 )%、(128±24)%;(143±32)%、(137±29)%对照组为100%,均显著高于对照组(P<0.05);低浓度组E2和P分泌量与对照组差异无显著性。随着细胞培养天数的增加 , GnRH-a中、高浓度组E2分泌量明显低于对照组,高浓度组E2分泌量与培养天数呈负相关 (r=-0.75,P<0.05)。结论:GnRH-a对黄素化颗粒细胞分 泌甾体激素功能的影响随着作用时间和浓度的不同而变化。%Objective: To observe the effect of gonadotrop in-r eleasing hormone agonist(GnRH-a) on steroidogenesis of cultured human luteinize d granulosa cells in vitro.Methods: Human luteinized granulosa c ells were cultured in serum-free McCoy'5a medium.After stimulating with various concentrations of GnRH-a for 2, 4 and 6 days.Estradoil (E2) and progesterone ( P ) levels in the media were measured by radioimmunoassay.Results: When stimulated with different concentrations (10.0ng/ml and 100.0ng/ml)of GnRH -a for 2 days, E2 and P levels produced by luteinized granulosa cells increase d and were significantly higher than those of control (P<0.05).In lower concentration group there were no significantly difference of E2 a nd P le vels when compared with control group.After stimulated with different concentrat ion of GnRH-a for 4 and 6 days,the E2 levels in media significantly decreased e xcept for the low concentration group.There was a significant positive correlati o n between the E2 level and days under the stimulation of high concentration Gn R H-a(r=-0.75,P<0.05).Conclusion

  10. Granulosa cells and retinoic acid co-treatment enrich potential germ cells from manually selected Oct4-EGFP expressing human embryonic stem cells.

    Science.gov (United States)

    Chen, Hsin-Fu; Jan, Pey-Shynan; Kuo, Hung-Chih; Wu, Fang-Chun; Lan, Chen-Wei; Huang, Mei-Chi; Chien, Chung-Liang; Ho, Hong-Nerng

    2014-09-01

    Differentiation of human embryonic stem (HES) cells to germ cells may become clinically useful in overcoming diseases related to germ-cell development. Niches were used to differentiate HES cell lines, NTU1 and H9 Oct4-enhanced green fluorescence protein (EGFP), including laminin, granulosa cell co-culture or conditioned medium, ovarian stromal cell co-culture or conditioned medium, retinoic acid, stem cell factor (SCF) and BMP4-BMP7-BMP8b treatment. Flow cytometry showed that granulosa cell co-culture (P cells expressing early germ cell marker stage-specific embryonic antigen 1(SSEA1); sorted SSEA1[+] cells did not express higher levels of germ cell gene VASA and GDF9. Manually collected H9 Oct4-EGFP[+] cells expressed significantly higher levels of VASA (P = 0.005) and GDF9 (P = 0.001). H9 Oct4-EGFP[+] cells developed to ovarian follicle-like structures after culture for 28 days but with low efficiency. Unlike SCF and BMP4, retinoic acid co-treatment enhanced VASA, GDF9 and SCP3 expression. A protocol is recommended to enrich differentiated HES cells with germ-cell potential by culture with granulosa cells, conditioned medium or retinoic acid, manual selection of Oct4-EGFP[+] cells, and analysis of VASA, GDF9 expression, or both.

  11. Exploratory metabolomic analyses reveal compounds correlated with lutein concentration in frontal cortex, hippocampus, and occipital cortex of human infant brain

    Science.gov (United States)

    Lutein is a dietary carotenoid well known for its role as an antioxidant in the macula and recent reports implicate a role for lutein in cognitive function. Lutein is the dominant carotenoid in both pediatric and geriatric brain tissue. In addition, cognitive function in older adults correlated with...

  12. Dissociation of human follicle-stimulating hormone. Comparison with luteinizing hormone.

    Science.gov (United States)

    Reichert, L E; Ramsey, R B

    1975-04-25

    Rat testis tissue receptor assays were utilized to study the kinetics of dissociation of human follicle-stimulating hormone (hFSH) and luteinizing hormone (hLH) under varying conditions of urea concentration and pH. In these competitive protein binding assays, 125I-hFSH and 125I-hLH were the radioligands and hormone dissociation was followed by a decrease in the ability of the dissociating hormone to inhibit uptake of the radioligand by tissue receptors. Rate data for dissociation of the gonadotropins were analyzed for quality of fit to first or second order integrated rate equations by nonlinear regression analysis. Treatment of hFSH with 4 M urea at pH 8 and 25 degrees for 22 hours did not result in significant dissociation, whereas in 8 M urea, over 90% dissociation was observed. The rate of dissociation of hFSH in 8 M urea was increased approximately 4-fold by raising the temperature from 25 to 37 degrees. Similar results were obtained when dissociation of hFSH was followed through use of an accepted whole animal bioassay for FSH, thus confirming the reliability of the tissue receptor assay for such dissociation studies. Kinetic studies showed that hFSH was undissociated by incubation in 6 M urea of pH 8 after 4 hours at 25 degrees. In contrast, hLH was 90% dissociated under similar conditions. This differential rate of inactivation of hLH allowed preparation of hFSH having significant reduced levels of contaminating LH activity, as determined by tissue receptor assays and by whole animal bioassays. Marked differences were noted in the rate of dissociation of hFSH and hLH under acid conditions. hFSH completely dissociated after approximately 2 min of incubation of pH 2 (25 degrees), and over 90% dissociated after 15 min of incubation at pH 3. In contrast, hLH was dissociated 60% after 20 min of incubation at pH 2 (25 degrees) and 40% dissociated after 60 min at pH 3. Neither hormone was significantly dissociated at pH 4.4 after 60 min, but hFSH showed a

  13. Data in support of FSH induction of IRS-2 in human granulosa cells: Mapping the transcription factor binding sites in human IRS-2 promoter

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    Surleen Kaur

    2016-03-01

    Full Text Available Insulin receptor substrate-2 (IRS-2 plays critical role in the regulation of various metabolic processes by insulin and IGF-1. The defects in its expression and/or function are linked to diseases like polycystic ovary syndrome (PCOS, insulin resistance and cancer. To predict the transcription factors (TFs responsible for the regulation of human IRS-2 gene expression, the transcription factor binding sites (TFBS and the corresponding TFs were investigated by analysis of IRS-2 promoter sequence using MatInspector Genomatix software (Cartharius et al., 2005 [1]. The ibid data is part of author׳s publication (Anjali et al., 2015 [2] that explains Follicle stimulating hormone (FSH mediated IRS-2 promoter activation in human granulosa cells and its importance in the pathophysiology of PCOS. Further analysis was carried out for binary interactions of TF regulatory genes in IRS-2 network using Cytoscape software tool and R-code. In this manuscript, we describe the methodology used for the identification of TFBSs in human IRS-2 promoter region and provide details on experimental procedures, analysis method, validation of data and also the raw files. The purpose of this article is to provide the data on all TFBSs in the promoter region of human IRS-2 gene as it has the potential for prediction of the regulation of IRS-2 gene in normal or diseased cells from patients with metabolic disorders and cancer.

  14. Effects of bioactive compounds from carrots (Daucus carota L.), polyacetylenes, beta-carotene and lutein on human lymphoid leukaemia cells.

    Science.gov (United States)

    Zaini, Rana G; Brandt, Kirsten; Clench, Malcolm R; Le Maitre, Christine L

    2012-07-01

    New therapies for leukaemia are urgently needed. Carrots have been suggested as a potential treatment for leukaemia in traditional medicine and have previously been studied in other contexts as potential sources of anticancer agents. Indicating that carrots may contain bioactive compounds, which may show potential in leukaemia therapies. This study investigated the effects of five fractions from carrot juice extract (CJE) on human lymphoid leukaemia cell lines, together with five purified bioactive compounds found in Daucus carota L, including: three polyacetylenes (falcarinol, falcarindiol and falcarindiol-3-acetate) and two carotenoids (beta-carotene and lutein). Their effects on induction of apoptosis using Annexin V/PI and Caspase 3 activity assays analysed via flow cytometry and inhibition of cellular proliferation using Cell Titer Glo assay and cell cycle analysis were investigated. Treatment of all three lymphoid leukaemia cell lines with the fraction from carrot extracts which contained polyacetylenes and carotenoids was significantly more cytotoxic than the 4 other fractions. Treatments with purified polyacetylenes also induced apoptosis in a dose and time responsive manner. Moreover, falcarinol and falcarindiol-3-acetate isolated from Daucus carota L were more cytotoxic than falcarindiol. In contrast, the carotenoids showed no significant effect on either apoptosis or cell proliferation in any of the cells investigated. This suggests that polyacetylenes rather than beta-carotene or lutein are the bioactive components found in Daucus carota L and could be useful in the development of new leukemic therapies. Here, for the first time, the cytotoxic effects of polyacetylenes have been shown to be exerted via induction of apoptosis and arrest of cell cycle.

  15. Brilliant cresyl blue staining does not present cytotoxic effects on human luteinized follicular cells, according to gene/protein expression, as well as to cytotoxicity tests.

    Science.gov (United States)

    Alcoba, Diego Duarte; Schneider, Júlia; Arruda, Letícia; Martiny, Patrícia Borba; Capp, Edison; von Eye Corleta, Helena; Brum, Ilma Simoni

    2017-03-01

    In vitro oocyte maturation presents many advantages and its success is related to the selection of fully grown oocytes. In animal models, staining of cumulus-oocyte complexes (COCs) with Brilliant Cresyl Blue (BCB) is widely used for this selection. However, a lack of information about the safety of BCB makes its applicability in humans questionable. Because granulosa and cumulus cells have a close relationship with the oocyte and taking into account that follicular cells are also exposed to BCB when this pre-selection method is applied, we aimed to evaluate the effects of BCB on human follicular cells exposed to BCB. Cytotoxicity tests (Sulforhodamine B and Neutral Red Uptake) and gene and protein expression of elements related to the cell cycle (BAX, BCL2, TP53 and CDKN1A), as well as to cell death and metabolism (GAPDH), glucose consumption, and estradiol and progesterone secretion, were examined in granulosa and cumulus cells with and without exposure to BCB. Regardless estradiol secretion and glucose consumption, all other evaluations presented similar results between control and treated (BCB) groups, independently of cell type. Therefore, our results demonstrate convincingly that BCB seems to be safe for use in humans and it should encourage future studies to evaluate the development of embryos derived from human oocytes selected by BCB staining. Copyright © 2016. Published by Elsevier Urban & Partner Sp. z o.o.

  16. Anti-Müllerian hormone inhibits growth of AMH type II receptor-positive human ovarian granulosa cell tumor cells by activating apoptosis.

    Science.gov (United States)

    Anttonen, Mikko; Färkkilä, Anniina; Tauriala, Hanna; Kauppinen, Marjut; Maclaughlin, David T; Unkila-Kallio, Leila; Bützow, Ralf; Heikinheimo, Markku

    2011-11-01

    Ovarian granulosa cell tumors (GCTs) are sex cord stromal tumors that constitute 3-5% of all ovarian cancers. GCTs usually present with an indolent course but there is a high risk of recurrence, which associates with increased mortality, and targeted treatments would be desirable. Anti-Müllerian hormone (AMH), a key factor regulating sexual differentiation of the reproductive organs, has been implicated as a growth inhibitor in ovarian cancer. GCTs and normal granulosa cells produce AMH, but its expression in large GCTs is usually downregulated. Further, as the lack of specific AMH-signaling pathway components leads to GCT development in mice, we hypothesized that AMH inhibits growth of GCTs. Utilizing a large panel of human GCT tissue samples, we found that AMH type I receptors (ALK2, ALK3 and ALK6) and type II receptor (AMHRII), as well as their downstream effectors Smad1/5, are expressed and active in GCTs. AMHRII expression was detected in the vast majority (96%) of GCTs and correlated with AMH mRNA and protein expression. AMH mRNA level was low in large GCTs, confirming previous findings on low-AMH protein expression in large human as well as mouse GCTs. To study the functional role of AMH in this peculiar ovarian cancer, we utilized a human GCT cell line (KGN) and 10 primary GCT cell cultures. We found that the AMH-Smad1/5-signaling pathway was active in these cells, and that exogenous AMH further activated Smad1/5 in KGN cells. Furthermore, AMH treatment reduced the number of KGN cells and primary GCT cells, with increasing amounts of AMH leading to augmented activation of caspase-3 and subsequent apoptosis. All in all, these data support the premise that AMH is a growth inhibitor of GCTs.

  17. [Advances in the researches of lutein and alzheimer's disease].

    Science.gov (United States)

    Xu, Xianrong; Lin, Xiaoming

    2015-05-01

    Lutein, a kind of oxycarotenoid, can pass the blood brain barrier and preferentially accumulate in the human brain, which is the most abundant carotenoid in human brain. Evidence from multiple studies suggested that lutein was closely related to age-related cognitive decline and risk of Alzheimer's disease (AD) in human. Dietary, plasma and brain concentrations of lutein were negatively associated with age-related cognitive decline. Lutein concentrations in plasma and brain were significantly lower in AD patients than those of health control. In human brain, lutein was the sole carotenoid which consistently associated with a range of cognitive function measures. In elderly women, lutein supplement can improve the cognitive function. In this article, we systematically reviewed the literature on the role of lutein in age-related cognitive decline and alzheimer's disease and its possible mechanisms. It may prove some benefit information for the advanced research and prevention of AD.

  18. Effects of BMAL1-SIRT1-positive cycle on estrogen synthesis in human ovarian granulosa cells: an implicative role of BMAL1 in PCOS.

    Science.gov (United States)

    Zhang, Jiaou; Liu, Jiansheng; Zhu, Kai; Hong, Yan; Sun, Yun; Zhao, Xiaoming; Du, Yanzhi; Chen, Zi-Jiang

    2016-08-01

    Brain and muscle ARNT-like protein 1 (BMAL1) is necessary for fertility and has been found to be essential to follicle growth and steroidogenesis. Sirtuin1 (SIRT1) has been reported to interact with BMAL1 and function in a circadian manner. Evidence has shown that SIRT1 regulates aromatase expression in estrogen-producing cells. We aimed to ascertain if there is a relationship between polycystic ovary syndrome (PCOS) and BMAL1, and whether and how BMAL1 takes part in estrogen synthesis in human granulosa cells (hGCs). Twenty-four women diagnosed with PCOS and 24 healthy individuals undergoing assisted reproduction were studied. BMAL1 expression in their granulosa cells (GCs) was observed by quantitative real-time polymerase chain reaction (qRT-PCR). The level of expression in the PCOS group was lower than that of the group without PCOS (p SIRT1 knock-down and, conversely, upregulated after overexpression treatments of these two genes in KGN cells. Both BMAL1 and SIRT1 had a mutually positive regulation, as did the phosphorylation of JNK. Furthermore, JNK overexpression increased estrogen synthesis activity and the expression levels of aromatase, BMAL1, and SIRT1. In KGN and hGCs, estrogen synthesis and aromatase expression were downregulated after treatment with JNK and SIRT1 inhibitors. In addition, BMAL1, SIRT1, and JNK expression levels were all downregulated. Our results demonstrate the effects of BMAL1 on estrogen synthesis in hGCs and suggest a BMAL1-SIRT1-JNK positive feedback cycle in this process, which points out an important role of BMAL1 in the development of PCOS.

  19. Effects of dietary supplementation with a combination of fish oil, bilberry extract, and lutein on subjective symptoms of asthenopia in humans.

    Science.gov (United States)

    Kawabata, Fuminori; Tsuji, Tomoko

    2011-12-01

    The aim of this study was to determine the effects of dietary supplementation with a combination of fish oil, bilberry extract, and lutein on subjective symptoms of asthenopia in humans by a double- blind, randomized, parallel-group, and placebo-controlled trial. In the Active group, eleven subjects ingested a supplement containing omega-3 fatty acid-rich fish oil (docosahexaenoic acid 783 mg/day, eicosapentaenoic acid 162 mg/day), bilberry extract (anthocyanidin 59 mg/day), and lutein (17.5 mg/day) in soft gel capsule form, every day for 4 weeks. In the Placebo group, nine subjects ingested placebo capsules. Before and after supplementation, subjects completed a questionnaire to determine their asthenopia symptoms and were also assessed for mental fatigue symptom by the visual analog scale (VAS) test. Asthenopia symptoms such as "stiff shoulder, low back pain", "frustration", "dry-eye", and "stuffy head" were improved in the Active group. Furthermore, a score of mental fatigue was improved after 4 weeks of supplementation, and no side effects were observed after the 4-week supplementation and a 2-week washout period in the Active group. These results suggest that dietary supplementation with the combination of omega-3 fatty acid-rich fish oil, bilberry extract, and lutein may safely improve subjective symptoms of asthenopia and mental fatigue in humans.

  20. Effect of permeation enhancer pretreatment on the iontophoresis of luteinizing hormone releasing hormone (LHRH) through human epidermal membrane (HEM).

    Science.gov (United States)

    Smyth, Hugh D C; Becket, Gordon; Mehta, Samir

    2002-05-01

    A 2 x 2 factorial design was performed to determine the effect of a permeation enhancer (oleic acid/propylene glycol), iontophoresis (2 V), and the combination of the two treatments on the permeation enhancement of a model peptide, LHRH (luteinizing hormone releasing hormone), through human epidermal membrane (HEM). In parallel studies, TEAB (tetraethylammonium bromide, a small ionic solute) and sucrose (an electroosmotic flow marker) were also investigated. Structural changes in the HEM were monitored via conductance measurements, differential scanning calorimetry (DSC), and infrared (IR) spectroscopy experiments. LHRH enhancement due to enhancer in combination with iontophoresis (I + E; 29.5 times passive permeability, P), was greater than during iontophoresis alone (I; 14.3) and enhancer treatment alone (E; 3.5). I + E had an additive effect of I and E, indicating the mechanisms of action of the individual enhancement strategies were likely to be located at different sites in the skin. Also, no synergistic enhancement was observed with I + E for either TEAB or sucrose. For TEAB, permeability enhancement due to I (approximately 1400) was much higher than that due to E (14.9), and no additive effect could be detected. For sucrose, E had no effect on either passive or iontophoretic permeability, eliminating the possibility that electroosmosis could explain increases in LHRH permeability. Evidence of synergy between E and I was found, with conductance measurements indicating that I + E synergistically increased the membrane permeability to conducting ions (Na+ and Cl-). It appears these pathways were not available for transport for the solutes used in the current study. DSC and IR investigations showed significant changes in stratum corneum lipid structure following E treatment but not following I. These findings probably arise from the localized action of iontophoresis compared with the bulk action of enhancer. In summary, increased LHRH delivery through HEM in

  1. Modulation of DNA-induced damage and repair capacity in humans after dietary intervention with lutein-enriched fermented milk.

    Directory of Open Access Journals (Sweden)

    Carmen Herrero-Barbudo

    Full Text Available Dietary factors provide protection against several forms of DNA damage. Additionally, consumer demand for natural products favours the development of bioactive food ingredients with health benefits. Lutein is a promising biologically active component in the food industry. The EFSA Panel on Dietetic Products, Nutrition and Allergies considers that protection from oxidative damage may be a beneficial physiological effect but that a cause and effect relationship has not been established. Thus, our aim was to evaluate the safety and potential functional effect of a lutein-enriched milk product using the Comet Assay in order to analyze the baseline, the induced DNA-damage and the repair capacity in the lymphocytes of 10 healthy donors before and after the intake of the mentioned product. Our data suggest that the regular consumption of lutein-enriched fermented milk results in a significant increase in serum lutein levels and this change is associated with an improvement in the resistance of DNA to damage and the capacity of DNA repair in lymphocytes. Our results also support the lack of a genotoxic effect at the doses supplied as well as the absence of interactions and side effects on other nutritional and biochemicals markers.

  2. The Impact of Polymeric Nanoencapsulation on the Bioavailability of Lutein

    Science.gov (United States)

    Kamil, Alison

    Lutein, a fat-soluble xanthophyll, contributes partially to the health benefits from consuming plant foods. Like all dietary carotenoids, lutein has a low bioavailability. In addition to increasing the intake of lutein-rich foods to enhance lutein status, delivery of lutein in polymeric nanoparticles (NP) presents a novel approach to enhancing lutein bioavailability. The overall research objective of this project was to investigate, in rats, the impact of nanoencapsulation using poly(lactic-co-glycolic acid) (PLGA) on the pharmacokinetics of lutein. We also used an in vitro cell culture approach utilizing human epithelial colorectal adenocarcinoma (Caco-2) cells grown in both conventional (CONV) and permeable support (PS) systems to investigate the impact of PLGA-NP on the absorption of lutein in intestinal cells. In chapter one, we compared the efficacy of lutein absorption in vitro using Caco-2 cells grown in both CONV and PS systems. We further examined the role of the micelle, the physiological vehicle for lutein within the small intestine, on its intestinal absorption in vitro compared to an organic solvent, ethanol, which is safe and consumed by humans. The finding from this study demonstrated that the CONV system displayed a larger efficacy of lutein uptake by Caco-2 cells. Further, in the PS system, micelle components appeared to facilitate more effective intestinal secretion of lutein. These findings suggest that lutein uptake by Caco-2 cells is subject to the influence of culturing system (CONV vs. PS) and delivery vehicle (ethanol vs. micelle). Chapter two examined the impact of PLGA-NP in rats on lutein pharmacokinetics in plasma and distribution in selected tissues as compared to free lutein. We also investigated the effect of nanoencapsulation on the absorption of lutein in intestinal cells compared to a more physiological vehicle, the micelle, using the PS method. In addition, we explored the need of additional micelles for the ultimate absorption of

  3. Induction of follicular luteinization by equine chorionic gonadotropin in cyclic guinea pigs.

    Science.gov (United States)

    Li, Jun-rong; Wang, Wei; Shi, Fang-xiong

    2015-12-01

    The effects of equine chorionic gonadotropin (eCG) on follicular development and ovulation in cyclic guinea pigs were investigated by histological and immunohistochemical analyses. Three groups of guinea pigs (n=12) were administrated subcutaneously with saline, 20 or 50 IU of eCG, respectively, on cyclic Day 12 (Day 1=vaginal openings). Ovaries were collected at 4 and 8 d after administration (6 animals per group each time). The eCG administration induced significant and distinct morphological changes in the ovaries, as it promoted the luteinization of granulosa cells, but not follicular development. In addition, proliferating cell nuclear antigen (PCNA) and steroidogenic acute regulatory protein (StAR) were immunolocalized specifically in luteinized follicles. Our experiments together indicate that eCG administration can induce follicular luteinization but not superovulation in guinea pigs. The eCG in cyclic guinea pigs functions similar to that of luteinizing hormone (LH), but not follicle-stimulating hormone (FSH).

  4. The potential role of granulosa cells in the maturation rate of immature human oocytes and embryo development: A co-culture study.

    Science.gov (United States)

    Jahromi, Bahia Namavar; Mosallanezhad, Zahra; Matloob, Najmeh; Davari, Maryam; Ghobadifar, Mohamed Amin

    2015-09-01

    In order to increase the number of mature oocytes usable for intracytoplasmic sperm injection (ICSI), we aimed to investigate the effect of co-culturing granulosa cells (GCs) on human oocyte maturation in vitro, the fertilization rate, and embryo development. A total of 133 immature oocytes were retrieved and were randomly divided into two groups; oocytes that were cultured with GCs (group A) and oocytes that were cultured without GCs (group B). After in vitro maturation, only oocytes that displayed metaphase II (MII) underwent the ICSI procedure. The maturation and fertilization rates were analyzed, as well as the frequency of embryo development. The mean age of the patients, their basal levels of follicle-stimulating hormone, and the number of oocytes recovered from the patients were all comparable between the two study groups. The number of oocytes that reached MII (mature oocytes) was 59 out of 70 (84.28%) in group A, compared to 41 out of 63 (65.07%) in group B (p=0.011). No significant difference between fertilization rates was found between the two study groups (p=0.702). The embryo development rate was higher in group A (33/59, 75%) than in group B (12/41, 42.85%; p=0.006). The proportion of highest-quality embryos and the blastocyst formation rate were significantly lower in group B than in group A (p=0.003 and p<0.001, respectively). The findings of the current study demonstrate that culturing immature human oocytes with GCs prior to ICSI improves the maturation rate and the likelihood of embryo development.

  5. Down-regulation of messenger ribonucleic acid encoding an importer of sulfoconjugated steroids during human chorionic gonadotropin-induced follicular luteinization in vivo.

    Science.gov (United States)

    Brown, Kristy A; Bouchard, Nadine; Lussier, Jacques G; Sirois, Jean

    2007-01-01

    Members of the organic anion transporting polypeptide (SLCO/OATP) superfamily are capable of importing anionic compounds across the lipid bilayer in a sodium-independent manner. Member 2B1 has been shown to transport few substrates, two of which are dihydroepiandrosterone-3-sulfate (DHEA-S) and estrone-3-sulfate. Steroid sulfatase (STS) catalyses the hydrolysis of these steroids into their unconjugated counterparts. The objective of this study was to investigate the regulation of SLCO2B1 and STS mRNAs during human chorionic gonadotropin (hCG)-induced ovulation/luteinization. The equine SLCO2B1 cDNA was cloned and shown to encode a 709-amino acid protein (OATP2B1) that is highly conserved when compared to mammalian orthologs. RT-PCR/Southern blot analyses were performed to study the regulation of SLCO2B1 and STS transcripts in equine preovulatory follicles isolated between 0 and 39h after hCG treatment. Results showed high levels of SLCO2B1 mRNA expression before hCG, with a marked decrease observed in follicles obtained 24-39h post-hCG (Pimport sulfoconjugated DHEA and estrogens, and that STS can remove the sulfonate moiety from these steroids, their down-regulation in luteinizing preovulatory follicles may provide an additional biochemical basis for the decrease in ovarian 17beta-estradiol biosynthesis after the LH surge.

  6. Regulatory role of BMP-9 in steroidogenesis by rat ovarian granulosa cells.

    Science.gov (United States)

    Hosoya, Takeshi; Otsuka, Fumio; Nakamura, Eri; Terasaka, Tomohiro; Inagaki, Kenichi; Tsukamoto-Yamauchi, Naoko; Hara, Takayuki; Toma, Kishio; Komatsubara, Motoshi; Makino, Hirofumi

    2015-03-01

    BMPs expressed in the ovary differentially regulate steroidogenesis by granulosa cells. BMP-9, a circulating BMP, is associated with cell proliferation, apoptosis and differentiation in various tissues. However, the effects of BMP-9 on ovarian function have yet to be elucidated. Here we investigated BMP-9 actions on steroidogenesis using rat primary granulosa cells. BMP-9 potently suppressed FSH-induced progesterone production, whereas it did not affect FSH-induced estradiol production by granulosa cells. The effects of BMP-9 on FSH-induced steroidogenesis were not influenced by the presence of oocytes. FSH-induced cAMP synthesis and FSH-induced mRNA expression of steroidogenic factors, including StAR, P450scc, 3βHSD2 and FSHR, were suppressed by treatment with BMP-9. BMP-9 mRNA expression was detected in granulosa cells but not in oocytes. BMP-9 readily activated Smad1/5/8 phosphorylation and Id-1 transcription in granulosa cells. Analysis using ALK inhibitors indicated that BMP-9 actions were mediated via type-I receptors other than ALK-2, -3 and -6. Furthermore, experiments using extracellular domains (ECDs) for BMP type-I and -II receptor constructs revealed that the effects of BMP-9 were reversed by ECDs for ALK-1 and BMPRII. Thus, the functional receptors for BMP-9 in granulosa cells were most likely to be the complex of ALK-1 and BMPRII. Collectively, the results of the present study showed that BMP-9 can affect luteinization and that there are two possible sources of BMP-9, serum and granulosa cells in the ovary.

  7. Dietary lutein absorption from marigold extract is rapid in BALB/c mice.

    Science.gov (United States)

    Park, J S; Chew, B P; Wong, T S

    1998-10-01

    Even though lutein can stimulate immunity and decrease cancer growth, no systematic studies are available on the uptake of lutein in mice. We studied the uptake of lutein in 8-wk-old female BALB/c mice fed a diet containing 0, 0.05, 0.1, 0.2 or 0.4% lutein. Mice were killed on d 0, 3, 7, 14, 21 and 28 (n = 6/period), and blood, spleen and liver were collected. Food intake and body, liver and spleen weights did not differ among treatment groups. Lutein + zeaxanthin were not detectable in the plasma, liver and spleen of unsupplemented mice. Mice fed lutein showed very rapid lutein + zeaxanthin absorption. On d 3, concentrations of plasma lutein + zeaxanthin had rapidly increased (P Plasma lutein + zeaxanthin concentrations did not differ among lutein-fed mice by d 7 (2.58 +/- 0.2 micromol/L). Even though maximal uptake of plasma lutein + zeaxanthin was observed by d 3, uptake of lutein + zeaxanthin by the liver and especially by the spleen generally continued to increase (P +/- 0.001 (spleen) and 0.71 +/- 0. 0002 (liver) nmol/g. Therefore, dietary lutein is readily absorbed into the plasma and taken up by liver and spleen of mice. Plasma lutein + zeaxanthin concentrations were higher than in human studies; however, mice were fed lutein at a level several hundredfold greater than in humans. The liver is a major storage organ for lutein + zeaxanthin in mice. Uptake of lutein + zeaxanthin by the spleen suggests a role for lutein in modulating immunity.

  8. Effect of the FSH receptor single nucleotide polymorphisms (FSHR 307/680) on the follicular fluid hormone profile and the granulosa cell gene expression in human small antral follicles

    DEFF Research Database (Denmark)

    Borgbo, Tanni Kjær; Jeppesen, J V; Lindgren, I

    2014-01-01

    ), by evaluating the hormone and gene expression profiles of human small antral follicles collected under physiological conditions in connection with fertility preservation. In total 69 women at various time during the menstrual cycle were included in this study. The intrafollicular hormone content of 179......The most pronounced effects of FSH signalling are potentially displayed in the follicle fluid, which acts as a reservoir for FSH-induced granulosa cell (GC) secreted hormones. This study investigates the effects of two common polymorphisms of FSHR, FSHR 307 (rs6165) and FSHR 680 (rs6166...... small antral follicles collected under physiological FSH conditions....

  9. Progestin and AdipoQ Receptor 7, Progesterone Membrane Receptor Component 1 (PGRMC1), and PGRMC2 and Their Role in Regulating Progesterone's Ability to Suppress Human Granulosa/Luteal Cells from Entering into the Cell Cycle.

    Science.gov (United States)

    Sueldo, Carolina; Liu, Xiufang; Peluso, John J

    2015-09-01

    The present studies were designed to determine the role of progesterone receptor membrane component 1 (PGRMC1), PGRMC2, progestin and adipoQ receptor 7 (PAQR7), and progesterone receptor (PGR) in mediating the antimitotic action of progesterone (P4) in human granulosa/luteal cells. For these studies granulosa/luteal cells of 10 women undergoing controlled ovarian hyperstimulation were isolated, maintained in culture, and depleted of PGRMC1, PGRMC2, PAQR7, or PGR by siRNA treatment. The rate of entry into the cell cycle was assessed using the FUCCI cell cycle sensor to determine the percentage of cells in the G1/S stage of the cell cycle. PGRMC1, PGRMC2, PAQR7, and PGR mRNA levels were assessed by real-time PCR and their interactions monitored by in situ proximity ligation assays (PLAs). These studies revealed that PGRMC1, PGRMC2, PAQR7, and PGR were expressed by granulosa/luteal cells from all patients, with PGRMC1 mRNA being most abundant, followed by PAQR7, PGRMC2, and PGR. However, their mRNA levels showed considerable patient variation. P4's ability to suppress entry into the cell cycle was dependent on PGRMC1, PGRMC2, and PAQR7 but not PGR. Moreover, PLAs indicated that PGRMC1, PGRMC2, and PAQR7 formed a complex within the cytoplasm. Based on these studies, it is proposed that these three P4 mediators form a complex within the cytoplasm that is required for P4's action. Moreover, P4's ability to regulate human follicle development may be dependent in part on the expression levels of each of these P4 mediators.

  10. Fractalkine restores the decreased expression of StAR and progesterone in granulosa cells from patients with polycystic ovary syndrome.

    Science.gov (United States)

    Huang, Shuo; Pang, Yanli; Yan, Jie; Lin, Shengli; Zhao, Yue; Lei, Li; Yan, Liying; Li, Rong; Ma, Caihong; Qiao, Jie

    2016-07-08

    Low progesterone levels are associated with luteal phase deficiency in women with polycystic ovary syndrome (PCOS). The mechanisms regulating progesterone biosynthesis in the granulosa cells from women with PCOS is largely unknown. Fractalkine is expressed in human ovaries, and is reported to regulate progesterone production in granulosa cells of healthy women. In the current study, we aimed to examine the role of fractalkine in women with PCOS. Reduced fractalkine levels were found in follicular fluid and granulosa cells, accompanied by decreased progesterone production and reduced steroidogenic acute regulatory protein (StAR) expression in the granulosa cells of patients with PCOS. Administration of fractalkine reversed the inhibition of progesterone and StAR expression. The mechanism mediating these effects may be associated with the inhibition of ERK activity in the granulosa cells from women with PCOS. Our findings revealed that fractalkine regulated steroidogenesis in follicular granulosa cells of women with PCOS.

  11. Fractalkine restores the decreased expression of StAR and progesterone in granulosa cells from patients with polycystic ovary syndrome

    Science.gov (United States)

    Huang, Shuo; Pang, Yanli; Yan, Jie; Lin, Shengli; Zhao, Yue; Lei, Li; Yan, Liying; Li, Rong; Ma, Caihong; Qiao, Jie

    2016-01-01

    Low progesterone levels are associated with luteal phase deficiency in women with polycystic ovary syndrome (PCOS). The mechanisms regulating progesterone biosynthesis in the granulosa cells from women with PCOS is largely unknown. Fractalkine is expressed in human ovaries, and is reported to regulate progesterone production in granulosa cells of healthy women. In the current study, we aimed to examine the role of fractalkine in women with PCOS. Reduced fractalkine levels were found in follicular fluid and granulosa cells, accompanied by decreased progesterone production and reduced steroidogenic acute regulatory protein (StAR) expression in the granulosa cells of patients with PCOS. Administration of fractalkine reversed the inhibition of progesterone and StAR expression. The mechanism mediating these effects may be associated with the inhibition of ERK activity in the granulosa cells from women with PCOS. Our findings revealed that fractalkine regulated steroidogenesis in follicular granulosa cells of women with PCOS. PMID:27386819

  12. Effect of the human follicle-stimulating hormone-binding inhibitor and its N-terminal fragment on follicle-stimulating hormone-induced progesterone secretion by granulosa cells in vitro

    Indian Academy of Sciences (India)

    Perinaaz R Wadia; Smita D Mahale; Tarala D Nandedkar

    2007-09-01

    Intrafollicular factors play an important role in folliculogenesis. The follicle-stimulating hormone (FSH)-binding inhibitor (FSHBI), purified by our laboratory from human ovarian follicular fluid, has been shown to suppress ovulation and induce follicular atresia/apoptosis in mice as well as impair fertility in marmosets, the new world monkeys. The octapeptide, a peptide corresponding to the N-terminal region of human FSHBI (hFSHBI), has been synthesized and also shows FSHBI activity in vitro. In the present study, we have attempted to identify the mechanism of action of the peptide in granulosa cell cultures. Rat granulosa cell cultures were treated with varying concentrations of the octapeptide or partially purified hFSHBI (gel chromatography fraction hGF2) in the presence or absence of human FSH (hFSH) and the amount of progesterone (P4) secreted in the culture supernatants after 3 h/48 h was estimated. Both hGF2 and the octapeptide failed to alter basal levels as well as 8-bromo cAMP-induced P4 production, while FSH-induced P4 secretion was inhibited in a dose-dependent manner. These studies reveal that the octapeptide, a fragment of FSHBI, and the native protein have similar activity in vitro and both compounds alter FSH action at the receptor level upstream of cAMP formation.

  13. Extraovarian granulosa cell tumor

    Directory of Open Access Journals (Sweden)

    Paul Prabir

    2009-04-01

    Full Text Available Extraovarian granulosa cell tumor (GCT is a very uncommon tumor, assumed to arise from the ectopic gonadal tissue along the embryonal route of the genital ridge. One such rare case of extraovarian GCT was encountered in a 58-year-old female who presented with a large intraabdominal lump. Computerized tomography revealed one large retroperitoneal mass measuring 15cm x 16cm and another mesenteric mass of 8cm x 5cm size. The patient had a history of hysterectomy with bilateral salpingooophorectomy 20 years ago for uterine leiomyoma. Ultrasonography-guided aspiration smears revealed cytological features suggestive of GCT. Histopathological examination of the excised masses showed features of adult-type GCT. Because metastatic epithelial tumors, particularly from the ovaries, may show identical morphology, immunostains for inhibin and epithelial membrane antigen (EMA were performed. The tumor showed positivity for inhibin while EMA was negative thus confirming the diagnosis of GCT. As this patient had no previous history of GCT and was oophorectomized 20 years ago, the tumor was considered as extraovarian. A diagnosis of extraovarian GCT should be carried out after excluding any previous history of GCT of the ovary. Immunostains help to differentiate GCTs from other neoplasms.

  14. The fungicide mancozeb induces toxic effects on mammalian granulosa cells

    Energy Technology Data Exchange (ETDEWEB)

    Paro, Rita [Department of Health Sciences, University of L' Aquila, Via Vetoio, L' Aquila (Italy); Tiboni, Gian Mario [Department of Medicine and Aging, Section of Reproductive Sciences, University “G. D' Annunzio”, Chieti-Pescara (Italy); Buccione, Roberto [Tumor Cell Invasion Laboratory, Consorzio Mario Negri Sud, Santa Maria Imbaro, Chieti (Italy); Rossi, Gianna; Cellini, Valerio [Department of Health Sciences, University of L' Aquila, Via Vetoio, L' Aquila (Italy); Canipari, Rita [Department of Anatomy, Histology, Forensic Medicine and Orthopedics, Section of Histology and Embryology, School of Pharmacy and Medicine, “Sapienza” University of Rome, Rome (Italy); Cecconi, Sandra, E-mail: sandra.cecconi@cc.univaq.it [Department of Health Sciences, University of L' Aquila, Via Vetoio, L' Aquila (Italy)

    2012-04-15

    The ethylene-bis-dithiocarbamate mancozeb is a widely used fungicide with low reported toxicity in mammals. In mice, mancozeb induces embryo apoptosis, affects oocyte meiotic spindle morphology and impairs fertilization rate even when used at very low concentrations. We evaluated the toxic effects of mancozeb on the mouse and human ovarian somatic granulosa cells. We examined parameters such as cell morphology, induction of apoptosis, and p53 expression levels. Mouse granulosa cells exposed to mancozeb underwent a time- and dose-dependent modification of their morphology, and acquired the ability to migrate but not to proliferate. The expression level of p53, in terms of mRNA and protein content, decreased significantly in comparison with unexposed cells, but no change in apoptosis was recorded. Toxic effects could be attributed, at least in part, to the presence of ethylenthiourea (ETU), the main mancozeb catabolite, which was found in culture medium. Human granulosa cells also showed dose-dependent morphological changes and reduced p53 expression levels after exposure to mancozeb. Altogether, these results indicate that mancozeb affects the somatic cells of the mammalian ovarian follicles by inducing a premalignant-like status, and that such damage occurs to the same extent in both mouse and human GC. These results further substantiate the concept that mancozeb should be regarded as a reproductive toxicant. Highlights: ► The fungicide mancozeb affects oocyte spindle morphology and fertilization rate. ► We investigated the toxic effects of mancozeb on mouse and human granulosa cells. ► Granulosa cells modify their morphology and expression level of p53. ► Mancozeb induces a premalignant-like status in exposed cells.

  15. Luteinizing hormone in testicular descent

    DEFF Research Database (Denmark)

    Toppari, Jorma; Kaleva, Marko M; Virtanen, Helena E

    2007-01-01

    . Insulin-like hormone-3 (INSL3) is suggested to be the main regulator of gubernacular development and therefore an apparent regulator of testicular descent. INSL3 production is also related to LH, and reduced INSL3 action is a possible cause for cryptorchidism. Cryptorchid boys have normal testosterone......A proper hypothalamus-pituitary-testis axis with normal androgen synthesis and action is a prerequisite for normal testicular descent. Various defects in this axis may result in cryptorchidism but endocrine abnormalities are rarely detected. Androgens regulate testicular descent but androgen action...... alone is not sufficient for normal testicular descent. The regulation of androgen production is influenced both by placental human chorionic gonadotropin (hCG) and pituitary luteinizing hormone (LH). There is evidence that the longer pregnancy continues, the more important role pituitary LH may have...

  16. Effects of the environmental contaminants DEHP and TCDD on estradiol synthesis and aryl hydrocarbon receptor and peroxisome proliferator-activated receptor signalling in the human granulosa cell line KGN.

    Science.gov (United States)

    Ernst, Jana; Jann, Johann-Christoph; Biemann, Ronald; Koch, Holger M; Fischer, Bernd

    2014-09-01

    Environmental contaminants binding to transcription factors, such as the aryl hydrocarbon receptor (AhR) and the alpha and gamma peroxisome proliferator-activated receptors (PPARs), contribute to adverse effects on the reproductive system. Expressing both the AhR and PPARs, the human granulosa cell line KGN offers the opportunity to investigate the regulatory mechanisms involved in receptor crosstalk, independent of overriding hormonal control. The aim of the present study was to investigate the impact of two environmental contaminants, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, an AhR ligand) and di-(2-ethylhexyl) phthalate (DEHP, a PPAR ligand), on gonadotrophin sensitivity and estrogen synthesis in KGN cells. Accumulation of the DEHP metabolite mono-(2-ethylhexyl) phthalate (MEHP) in DEHP-exposed cells was measured by high-performance liquid chromatography mass spectrometry, thereby demonstrating DEHP metabolism to MEHP by KGN cells. By employing TCDD ( an AhR agonist), rosiglitazone (a PPARgamma agonist) or bezafibrate (a PPARalpha agonist), the presence of a functional AhR and PPAR cascade was confirmed in KGN cells. Cytotoxicity testing revealed no effect on KGN cell proliferation for the concentrations of TCDD and DEHP used in the current study. FSH-stimulated cells were exposed to TCDD, DEHP or a mix of both and estradiol synthesis was measured by enzyme-linked immunosorbent assay and gene expression by quantitative RT-PCR. Exposure decreased estradiol synthesis (TCDD, DEHP, mix) and reduced the mRNA expression of CYP19 aromatase (DEHP, mix) and FSHR (DEHP). DEHP induced the expression of the alpha and gamma PPARs and AhR, an effect which was inhibited by selective PPAR antagonists. Studies in the human granulosa cell line KGN show that the action of endocrine-disrupting chemicals may be due to a direct activation of AhR, for example by TCDD, and by a transactivation via PPARs, for example by DEHP, inducing subsequent transcriptional changes with a broad

  17. Two natural products, trans-phytol and (22E)-ergosta-6,9,22-triene-3β,5α,8α-triol, inhibit the biosynthesis of estrogen in human ovarian granulosa cells by aromatase (CYP19)

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Jiajia [Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu (China); Yuan, Yun [Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu (China); School of Life Science and Engineering, Southwest University of Science and Technology, Mianyang (China); Lu, Danfeng; Du, Baowen [Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu (China); Xiong, Liang; Shi, Jiangong [State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing (China); Yang, Lijuan [Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu (China); Liu, Wanli [MOE Key Laboratory of Protein Science, School of Life Sciences, Tsinghua University, Beijing 100084 (China); Yuan, Xiaohong [School of Life Science and Engineering, Southwest University of Science and Technology, Mianyang (China); Zhang, Guolin, E-mail: zhanggl@cib.ac.cn [Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu (China); Chinese Academy of Sciences Sichuan Translational Medicine Research Hospital, Chengdu (China); Wang, Fei, E-mail: wangfei@cib.ac.cn [Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu (China); Chinese Academy of Sciences Sichuan Translational Medicine Research Hospital, Chengdu (China)

    2014-08-15

    Aromatase is the only enzyme in vertebrates to catalyze the biosynthesis of estrogens. Although inhibitors of aromatase have been developed for the treatment of estrogen-dependent breast cancer, the whole-body inhibition of aromatase causes severe adverse effects. Thus, tissue-selective aromatase inhibitors are important for the treatment of estrogen-dependent cancers. In this study, 63 natural products with diverse structures were examined for their effects on estrogen biosynthesis in human ovarian granulosa-like KGN cells. Two compounds—trans-phytol (SA-20) and (22E)-ergosta-6,9,22-triene-3β,5α,8α-triol (SA-48)—were found to potently inhibit estrogen biosynthesis (IC{sub 50}: 1 μM and 0.5 μM, respectively). Both compounds decreased aromatase mRNA and protein expression levels in KGN cells, but had no effect on the aromatase catalytic activity in aromatase-overexpressing HEK293A cells and recombinant expressed aromatase. The two compounds decreased the expression of aromatase promoter I.3/II. Neither compound affected intracellular cyclic AMP (cAMP) levels, but they inhibited the phosphorylation or protein expression of cAMP response element-binding protein (CREB). The effects of these two compounds on extracellular regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinases (MAPKs), and AKT/phosphoinositide 3-kinase (PI3K) pathway were examined. Inhibition of p38 MAPK could be the mechanism underpinning the actions of these compounds. Our results suggests that natural products structurally similar to SA-20 and SA-48 may be a new source of tissue-selective aromatase modulators, and that p38 MAPK is important in the basal control of aromatase in ovarian granulosa cells. SA-20 and SA-48 warrant further investigation as new pharmaceutical tools for the prevention and treatment of estrogen-dependent cancers. - Highlights: • Two natural products inhibited estrogen biosynthesis in human ovarian granulosa cells. • They

  18. Enhanced Anti-Tumoral Activity of Methotrexate-Human Serum Albumin Conjugated Nanoparticles by Targeting with Luteinizing Hormone-Releasing Hormone (LHRH) Peptide

    Science.gov (United States)

    Taheri, Azade; Dinarvand, Rassoul; Atyabi, Fatemeh; Ahadi, Fatemeh; Nouri, Farank Salman; Ghahremani, Mohammad Hossein; Ostad, Seyed Nasser; Borougeni, Atefeh Taheri; Mansoori, Pooria

    2011-01-01

    Active targeting could increase the efficacy of anticancer drugs. Methotrexate-human serum albumin (MTX-HSA) conjugates, functionalized by luteinizing hormone-releasing hormone (LHRH) as targeting moieties, with the aim of specifically targeting the cancer cells, were prepared. Owing to the high expression of LHRH receptors in many cancer cells as compared to normal cells, LHRH was used as the targeting ligand in this study. LHRH was conjugated to MTX-HSA nanoparticles via a cross-linker. Three types of LHRH targeted nanoparticles with a mean particle size between 120–138 nm were prepared. The cytotoxicity of LHRH targeted and non-targeted nanoparticles were determined on the LHRH positive and negative cell lines. The internalization of the targeted and non-targeted nanoparticles in LHRH receptor positive and negative cells was investigated using flow cytometry analysis and fluorescence microscopy. The cytotoxicity of the LHRH targeted nanoparticles on the LHRH receptor positive cells were significantly more than non-targeted nanoparticles. LHRH targeted nanoparticles were also internalized by LHRH receptor positive cells significantly more than non-targeted nanoparticles. There were no significant differences between the uptake of targeted and non-targeted nanoparticles to the LHRH receptor negative cells. The active targeting procedure using LHRH targeted MTX-HSA nanoparticles could increase the anti-tumoral activity of MTX. PMID:21845098

  19. Enhanced Anti-Tumoral Activity of Methotrexate-Human Serum Albumin Conjugated Nanoparticles by Targeting with Luteinizing Hormone-Releasing Hormone (LHRH Peptide

    Directory of Open Access Journals (Sweden)

    Pooria Mansoori

    2011-07-01

    Full Text Available Active targeting could increase the efficacy of anticancer drugs. Methotrexate-human serum albumin (MTX-HSA conjugates, functionalized by luteinizing hormone-releasing hormone (LHRH as targeting moieties, with the aim of specifically targeting the cancer cells, were prepared. Owing to the high expression of LHRH receptors in many cancer cells as compared to normal cells, LHRH was used as the targeting ligand in this study. LHRH was conjugated to MTX-HSA nanoparticles via a cross-linker. Three types of LHRH targeted nanoparticles with a mean particle size between 120–138 nm were prepared. The cytotoxicity of LHRH targeted and non-targeted nanoparticles were determined on the LHRH positive and negative cell lines. The internalization of the targeted and non-targeted nanoparticles in LHRH receptor positive and negative cells was investigated using flow cytometry analysis and fluorescence microscopy. The cytotoxicity of the LHRH targeted nanoparticles on the LHRH receptor positive cells were significantly more than non-targeted nanoparticles. LHRH targeted nanoparticles were also internalized by LHRH receptor positive cells significantly more than non-targeted nanoparticles. There were no significant differences between the uptake of targeted and non-targeted nanoparticles to the LHRH receptor negative cells. The active targeting procedure using LHRH targeted MTX-HSA nanoparticles could increase the anti-tumoral activity of MTX.

  20. Role of leptin receptors in granulosa cells during ovulation.

    Science.gov (United States)

    Dupuis, Lisa; Schuermann, Yasmin; Cohen, Tamara; Siddappa, Dayananda; Kalaiselvanraja, Anitha; Pansera, Melissa; Bordignon, Vilceu; Duggavathi, Raj

    2014-02-01

    Leptin is an important hormone influencing reproductive function. However, the mechanisms underpinning the role of leptin in the regulation of reproduction remain to be completely deciphered. In this study, our objective is to understand the mechanisms regulating the expression of leptin receptor (Lepr) and its role in ovarian granulosa cells during ovulation. First, granulosa cells were collected from superovulated mice to profile mRNA expression of Lepr isoforms (LeprA and LeprB) throughout follicular development. Expression of LeprA and LeprB was dramatically induced in the granulosa cells of ovulating follicles at 4 h after human chorionic gonadotropin (hCG) treatment. Relative abundance of both mRNA and protein of CCAAT/enhancer-binding protein β (Cebpβ) increased in granulosa cells from 1 to 7 h post-hCG. Furthermore, chromatin immunoprecipitation assay confirmed the recruitment of Cebpβ to Lepr promoter. Thus, hCG-induced transcription of Lepr appears to be regulated by Cebpβ, which led us to hypothesise that Lepr may play a role during ovulation. To test this hypothesis, we used a recently developed pegylated superactive mouse leptin antagonist (PEG-SMLA) to inhibit Lepr signalling during ovulation. I.p. administration of PEG-SMLA (10 μg/g) to superovulated mice reduced ovulation rate by 65% compared with control treatment. Although the maturation stage of the ovulated oocytes remained unaltered, ovulation genes Ptgs2 and Has2 were downregulated in PEG-SMLA-treated mice compared with control mice. These results demonstrate that Lepr is dramatically induced in the granulosa cells of ovulating follicles and this induction of Lepr expression requires the transcription factor Cebpβ. Lepr plays a critical role in the process of ovulation by regulating, at least in part, the expression of the important genes involved in the preovulatory maturation of follicles.

  1. Incorporation of lutein and docosahexaenoic acid from dietary microalgae into the retina in quail.

    Science.gov (United States)

    Schnebelen-Berthier, Coralie; Acar, Niyazi; Pouillart, Philippe; Thabuis, Clementine; Rodriguez, Bertrand; Depeint, Flore; Clerc, Elise; Mathiaud, Adeline; Bourdillon, Anne; Baert, Blandine; Bretillon, Lionel; Lecerf, Jean-Michel

    2015-03-01

    Lutein and docosahexaenoic acid (DHA) are associated with the prevention of age-related macular degeneration (AMD). Since microalgae are potent natural sources of these nutrients, their nutritional value should be evaluated based on the bioavailability of lutein and DHA for the retina via the plasmatic compartment. In this study, quail were fed for 5 months either with a diet supplemented or deprived with microalgae rich in lutein and DHA. In the microalgae-fed group, the retinal concentrations of lutein and zeaxanthin gradually increased whereas in plasma, these compounds started to increase from the first month of supplementation. We also observed a significant increase in retinal and plasmatic levels of DHA in the microalgae-fed group. In conclusion, the plasmatic and retinal contents of lutein and DHA were significantly increased in quail fed with lutein- and DHA-rich microalgae. Food fortification with microalgae may be an innovative way to increase lutein and DHA consumption in humans.

  2. Why lutein is important for the eye and the brain

    Directory of Open Access Journals (Sweden)

    Ramirez Maria

    2016-01-01

    Full Text Available Lutein and zeaxanthin are carotenoids that accumulate in the macula. The macula is a yellow spot near the center of the retina that is responsible of high resolution vision. Macular pigment acts as a natural blue light filter and protects the eye from damage. Macular pigment optical density (MPOD increases with lutein administration and is related to visual function and to the prevention of age-related macular degeneration. MOPD can be measured non-invasively and has been related to better cognitive performance. Moreover, compositional analyses of centenarian brains have shown that lutein is the main carotenoid in the brain although not in plasma, indicating a preferential accumulation in neural tissues, and that carotenoids status is correlated with some functional outcomes. Carotenoids are present in human milk with higher concentration in colostrum than in transitional and mature milk. Formula fed-infants have less plasma lutein concentration than breast fed infants. Analyses of brain from infants who died during the first year of life showed that lutein is also the predominant carotenoid of brain. Studies in non-human primates revealed that carotenoids are determinant in the formation of the retinal epithelia. In vitro studies showed that lutein stimulates the differentiation of human stem cells to neural progenitor cells. All this findings together, mostly the presence of lutein in breast milk, plasma concentration in breast-fed infants vs. formula fed infants, preferential accumulation in the brain and evidences of influence on the retina and the functionality of the brain signal the importance of the role of lutein and zeaxanthin on visual maturation and brain development.

  3. Lutein as protective agent against neonatal oxidative stress

    Directory of Open Access Journals (Sweden)

    Giuseppe Buonocore

    2014-06-01

    Full Text Available Free radicals (FR are important for a correct development of neonatal organs and tissues. However, newborn and fetus have profoundly impaired antioxidant system. In these subjects, oxidative stress (OS may be detrimental by activating deleterious cellular processes. Decreasing FR and restoring oxidative imbalance certainly appear to be beneficial in perinatal period. Among the therapeutic antioxidant approaches in newborns, lutein, a compound belonging to the xanthophyll family of carotenoids, is one of the emerging strategies. Humans cannot synthesize lutein, hence the intake primarily depends on diet. In the neonatal period, fresh, non-processed human milk is the main dietary source of lutein, while infant formula is lacking it. Lutein has antioxidant and anti-inflammatory properties. Lutein supplementation in human newborns during the first days of life has been demonstrated to decrease plasma biomarkers of OS and increase antioxidant capacities. Numerous experimental study have demonstrated that lutein effectively neutralizes oxidants and modulates inflammatory processes, showing particular protective effects on macula and photoreceptors against phototoxicity and oxidative injury. Only few clinical studies evaluated the effectiveness of lutein in reducing preterm and term infant morbidity, reporting no definitive results. The challenge for the future is to better clarify the timing, the optimal dose and the duration of lutein intervention in perinatal period and to verify its impact on infants’ health. Proceedings of the 10th International Workshop on Neonatology · Cagliari (Italy · October 22nd-25th, 2014 · The last ten years, the next ten years in Neonatology Guest Editors: Vassilios Fanos, Michele Mussap, Gavino Faa, Apostolos Papageorgiou

  4. Heterologous radioimmunoassay for llama and alpaca luteinizing hormone with a monoclonal antibody, and equine standard and a human tracer

    Energy Technology Data Exchange (ETDEWEB)

    Aba, M.A.; Forsberg, M. [Swedish Univ. of Agricultural Sciences, Dept. of Clinical Chemistry, Faculty of Veterinary medicine, Uppsala (Sweden)

    1995-12-31

    A radioimmunoassay for llama and alpaca LH was developed using a human I{sup 125}LH tracer from a commercial kit, equine LH diluted in human LH free serum as standard, and amonoclonal antibody (518B7) specific for LH but with low species specificity. A 60-min delay in the addition of the tracer and overnight incubation gave a sensitivity of 0.8 {mu}3g L{sup -1}. The intra-assay coefficient of variation was 37% at 1 {mu}g L{sup -1}, declined to 15% at 4 {mu}g L{sup -1} and was below 6% for concentrations up to 32 {mu}g L{sup -1}. The inter-assay coefficients of variation for 3 control samples were 20% (2.8 {mu}g L{sup -1}), 16% (7.1 {mu}g L{sup -1}) and 9.8% (19 {mu}g L{sup -1}). In an attempt to increase sensitivity, all tubes were preincubated for 4 h at room temperature before adding the tracer, and the sample volume was increased from 50 {mu}L to 100 {mu}L (in the standard curve the increased volume was compensated for by human LH free serum). With this protocol, the assay sensitivity was 0,5 {mu}g L{sup -1}. The assay was validated clinically and demonstrated increased concentrations of LH after mating in llamas and alpacas. Furthermore, the assay was used to monitor LH responses to a single dose of GnRH in llamas (adult males and females at different ages). (au) 9 refs.

  5. Genotoxicity of Superparamagnetic Iron Oxide Nanoparticles in Granulosa Cells

    Directory of Open Access Journals (Sweden)

    Marina Pöttler

    2015-11-01

    Full Text Available Nanoparticles that are aimed at targeting cancer cells, but sparing healthy tissue provide an attractive platform of implementation for hyperthermia or as carriers of chemotherapeutics. According to the literature, diverse effects of nanoparticles relating to mammalian reproductive tissue are described. To address the impact of nanoparticles on cyto- and genotoxicity concerning the reproductive system, we examined the effect of superparamagnetic iron oxide nanoparticles (SPIONs on granulosa cells, which are very important for ovarian function and female fertility. Human granulosa cells (HLG-5 were treated with SPIONs, either coated with lauric acid (SEONLA only, or additionally with a protein corona of bovine serum albumin (BSA; SEONLA-BSA, or with dextran (SEONDEX. Both micronuclei testing and the detection of γH2A.X revealed no genotoxic effects of SEONLA-BSA, SEONDEX or SEONLA. Thus, it was demonstrated that different coatings of SPIONs improve biocompatibility, especially in terms of genotoxicity towards cells of the reproductive system.

  6. Luteinizing hormone (LH) blood test

    Science.gov (United States)

    ICSH - blood test; Luteinizing hormone - blood test; Interstitial cell stimulating hormone - blood test ... to temporarily stop medicines that may affect the test results. Be sure to tell your provider about ...

  7. Lutropin alpha, recombinant human luteinizing hormone, for the stimulation of follicular development in profoundly LH-deficient hypogonadotropic hypogonadal women: a review

    Directory of Open Access Journals (Sweden)

    Bernd Th Krause

    2009-06-01

    Full Text Available Bernd Th Krause1, Ralf Ohlinger2, Annette Haase31Center for Endocrinology and Reproductive Medicine, MVZ Uhlandstr, Berlin, Germany; 2Ernst-Moritz-Arndt-University, Department of Gynecology and Obstetrics, Greifswald, Germany; 3Uhlandstr. 162, 10719 BerlinAbstract: Hypogonadotropic hypogonadism is defined as a medical condition with low or undetectable gonadotropin secretion, associated with a complete arrest of follicular growth and very low estradiol. The main cause can be traced back to an irregular or absent hypothalamic GnRH secretion, whereas only a minority suffers from a pituitary disorder. The choice of treatment to reverse this situation is a pulsatile GnRH application or a direct ovarian stimulation using gonadotropin injections. The goal is to achieve a proper ovarian function in these cases for a short time to allow ovulation and chance of pregnancy. Since the pulsatile GnRH treatment lost its former importance, several gonadotropins are in use to stimulate follicular growth, such as urine-derived human menopausal gonadotropin, highly purified follicle stimulating hormone (FSH or recombinant FSH, all with different success. The introduction of recombinant luteinizing hormone (LH and FSH provided an opportunity to investigate the distinct influences of LH and FSH alone and in combination on follicular growth in monofollicular ovulation induction cycles, and additionally on oocyte maturation, fertilization competence of the oocyte and embryo quality in downregulated IVF patients. Whereas FSH was known to be indispensable for normal follicular growth, the role of LH remained questionable. Downregulated IVF patients with this short-term gonadotropin depletion displayed no advance in stimulation success with the use of recombinant LH. Patients with hypogonadotropic hypogonadism undergoing monofollicular stimulation for ovulation induction showed clearly a specific role and need for both hormones in normal follicular growth. Therefore, a

  8. Data in support of FSH induction of IRS-2 in human granulosa cells: Mapping the transcription factor binding sites in human IRS-2 promoter

    OpenAIRE

    Surleen Kaur; Anjali, G.; Priya Bhardwaj; Jyoti Taneja; Rita Singh

    2015-01-01

    Insulin receptor substrate-2 (IRS-2) plays critical role in the regulation of various metabolic processes by insulin and IGF-1. The defects in its expression and/or function are linked to diseases like polycystic ovary syndrome (PCOS), insulin resistance and cancer. To predict the transcription factors (TFs) responsible for the regulation of human IRS-2 gene expression, the transcription factor binding sites (TFBS) and the corresponding TFs were investigated by analysis of IRS-2 promoter sequ...

  9. Cytotoxicity and mitogenicity assays with real-time and label-free monitoring of human granulosa cells with an impedance-based signal processing technology intergrating micro-electronics and cell biology.

    Science.gov (United States)

    Oktem, Ozgur; Bildik, Gamze; Senbabaoglu, Filiz; Lack, Nathan A; Akin, Nazli; Yakar, Feridun; Urman, Defne; Guzel, Yilmaz; Balaban, Basak; Iwase, Akira; Urman, Bulent

    2016-04-01

    A recently developed technology (xCelligence) integrating micro-electronics and cell biology allows real-time, uninterrupted and quantitative analysis of cell proliferation, viability and cytotoxicity by measuring the electrical impedance of the cell population in the wells without using any labeling agent. In this study we investigated if this system is a suitable model to analyze the effects of mitogenic (FSH) and cytotoxic (chemotherapy) agents with different toxicity profiles on human granulosa cells in comparison to conventional methods of assessing cell viability, DNA damage, apoptosis and steroidogenesis. The system generated the real-time growth curves of the cells, and determined their doubling times, mean cell indices and generated dose-response curves after exposure to cytotoxic and mitogenic stimuli. It accurately predicted the gonadotoxicity of the drugs and distinguished less toxic agents (5-FU and paclitaxel) from more toxic ones (cisplatin and cyclophosphamide). This platform can be a useful tool for specific end-point assays in reproductive toxicology. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. The Role of Lutein in Eye-Related Disease

    Directory of Open Access Journals (Sweden)

    Kakarla V. Chalam

    2013-05-01

    Full Text Available The lens and retina of the human eye are exposed constantly to light and oxygen. In situ phototransduction and oxidative phosphorylation within photoreceptors produces a high level of phototoxic and oxidative related stress. Within the eye, the carotenoids lutein and zeaxanthin are present in high concentrations in contrast to other human tissues. We discuss the role of lutein and zeaxanthin in ameliorating light and oxygen damage, and preventing age-related cellular and tissue deterioration in the eye. Epidemiologic research shows an inverse association between levels of lutein and zeaxanthin in eye tissues and age related degenerative diseases such as macular degeneration (AMD and cataracts. We examine the role of these carotenoids as blockers of blue-light damage and quenchers of oxygen free radicals. This article provides a review of possible mechanisms of lutein action at a cellular and molecular level. Our review offers insight into current clinical trials and experimental animal studies involving lutein, and possible role of nutritional intervention in common ocular diseases that cause blindness.

  11. The Role of Lutein in Eye-Related Disease

    Science.gov (United States)

    Koushan, Keyvan; Rusovici, Raluca; Li, Wenhua; Ferguson, Lee R.; Chalam, Kakarla V.

    2013-01-01

    The lens and retina of the human eye are exposed constantly to light and oxygen. In situ phototransduction and oxidative phosphorylation within photoreceptors produces a high level of phototoxic and oxidative related stress. Within the eye, the carotenoids lutein and zeaxanthin are present in high concentrations in contrast to other human tissues. We discuss the role of lutein and zeaxanthin in ameliorating light and oxygen damage, and preventing age-related cellular and tissue deterioration in the eye. Epidemiologic research shows an inverse association between levels of lutein and zeaxanthin in eye tissues and age related degenerative diseases such as macular degeneration (AMD) and cataracts. We examine the role of these carotenoids as blockers of blue-light damage and quenchers of oxygen free radicals. This article provides a review of possible mechanisms of lutein action at a cellular and molecular level. Our review offers insight into current clinical trials and experimental animal studies involving lutein, and possible role of nutritional intervention in common ocular diseases that cause blindness. PMID:23698168

  12. Phase II enzyme induction by a carotenoid, lutein, in a PC12D neuronal cell line

    Energy Technology Data Exchange (ETDEWEB)

    Miyake, Seiji [Laboratory of Retinal Cell Biology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582 (Japan); Department of Ophthalmology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582 (Japan); Wakasa Seikatsu Co., Ltd., 134 Chudoujiminami-cho, Shimogyo-ku, Kyoto 600-8813 (Japan); Kobayashi, Saori [Wakasa Seikatsu Co., Ltd., 134 Chudoujiminami-cho, Shimogyo-ku, Kyoto 600-8813 (Japan); Tsubota, Kazuo [Laboratory of Retinal Cell Biology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582 (Japan); Ozawa, Yoko, E-mail: ozawa@a5.keio.jp [Laboratory of Retinal Cell Biology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582 (Japan); Department of Ophthalmology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582 (Japan)

    2014-04-04

    Highlights: • Lutein reduced ROS levels in a PC12D neuronal cell line. • Lutein induced mRNAs of phase II antioxidative enzymes in PC12D neuronal cells. • Lutein increased protein levels of HO-1, SOD2, and NQO-1 in PC12D neuronal cells. • Lutein had no effect on intranuclear Nrf2 levels in PC12D neuronal cells. • Lutein did not activate potential upstream Nrf2 nuclear translocation pathways. - Abstract: The mechanism by which lutein, a carotenoid, acts as an antioxidant in retinal cells is still not fully understood. Here, lutein treatment of a neuronal cell line (PC12D) immediately resulted in reduced intracellular ROS levels, implying that it has a direct role in ROS scavenging. Significantly, lutein treatment also induced phase II antioxidative enzyme expression, probably via a nuclear factor-like 2 (Nrf2) independent pathway. This latter mechanism could explain why lutein acts diversely to protect against oxidative/cytotoxic stress, and why it is physiologically involved in the human neural tissue, such as the retina.

  13. Defective CFTR-regulated granulosa cell proliferation in polycystic ovarian syndrome.

    Science.gov (United States)

    Chen, Hui; Guo, Jing Hui; Zhang, Xiao Hu; Chan, Hsiao Chang

    2015-05-01

    Polycystic ovarian syndrome (PCOS) is one of the most frequent causes of female infertility, featured by abnormal hormone profile, chronic oligo/anovulation, and presence of multiple cystic follicles in the ovary. However, the mechanism underlying the abnormal folliculogenesis remains obscure. We have previously demonstrated that CFTR, a cAMP-dependent Cl(-) and HCO3 (-) conducting anion channel, is expressed in the granulosa cells and its expression is downregulated in PCOS rat models and human patients. In this study, we aimed to investigate the possible involvement of downregulation of CFTR in the impaired follicle development in PCOS using two rat PCOS models and primary culture of granulosa cells. Our results indicated that the downregulation of CFTR in the cystic follicles was accompanied by reduced expression of proliferating cell nuclear antigen (PCNA), in rat PCOS models. In addition, knockdown or inhibition of CFTR in granulosa cell culture resulted in reduced cell viability and downregulation of PCNA. We further demonstrated that CFTR regulated both basal and FSH-stimulated granulosa cell proliferation through the HCO3 (-)/sAC/PKA pathway leading to ERK phosphorylation and its downstream target cyclin D2 (Ccnd2) upregulation. Reduced ERK phosphorylation and CCND2 were found in ovaries of rat PCOS model compared with the control. This study suggests that CFTR is required for normal follicle development and that its downregulation in PCOS may inhibit granulosa cell proliferation, resulting in abnormal follicle development in PCOS.

  14. Bioavailability and biodistribution of nanodelivered lutein.

    Science.gov (United States)

    Kamil, Alison; Smith, Donald E; Blumberg, Jeffrey B; Astete, Carlos; Sabliov, Cristina; Oliver Chen, C-Y

    2016-02-01

    The aim of the study was to evaluate the ability of poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NP) to enhance lutein bioavailability. The bioavailability of free lutein and PLGA-NP lutein in rats was assessed by determining plasma pharmacokinetics and deposition in selected tissues. Lutein uptake and secretion was also assessed in Caco-2 cells. Compared to free lutein, PLGA-NP increased the maximal plasma concentration (Cmax) and area under the time-concentration curve in rats by 54.5- and 77.6-fold, respectively, while promoting tissue accumulation in the mesenteric fat and spleen. In comparison with micellized lutein, PLGA-NP lutein improved the Cmax in rat plasma by 15.6-fold and in selected tissues by ⩾ 3.8-fold. In contrast, PLGA-NP lutein had a lower uptake and secretion of lutein in Caco-2 cells by 10.0- and 50.5-fold, respectively, compared to micellized lutein. In conclusion, delivery of lutein with polymeric NP may be an approach to improve the bioavailability of lutein in vivo.

  15. Insights into granulosa cell tumors using spontaneous or genetically engineered mouse models.

    Science.gov (United States)

    Kim, So-Youn

    2016-03-01

    Granulosa cell tumors (GCTs) are rare sex cord-stromal tumors that have been studied for decades. However, their infrequency has delayed efforts to research their etiology. Recently, mutations in human GCTs have been discovered, which has led to further research aimed at determining the molecular mechanisms underlying the disease. Mouse models have been important tools for studying GCTs, and have provided means to develop and improve diagnostics and therapeutics. Thus far, several genetically modified mouse models, along with one spontaneous mouse model, have been reported. This review summarizes the phenotypes of these mouse models and their applicability in elucidating the mechanisms of granulosa cell tumor development.

  16. Gonadotropin regulation of the rat proopiomelanocortin promoter: characterization by transfection of primary ovarian granulosa cells.

    Science.gov (United States)

    Young, S L; Nielsen, C P; Lundblad, J R; Roberts, J L; Melner, M H

    1989-01-01

    To characterize the transcriptional effects of human (h)FSH and hCG on the POMC gene, primary rat granulosa cells were transiently transfected with a chloramphenicol acetyltransferase (CAT) reporter plasmid under the control of the POMC promoter and 5' region. POMC-CAT contains a fragment of the rat POMC gene, extending from nucleotide -704 to nucleotide +63, fused to the CAT gene. Treatment of POMC-CAT-transfected cells with either hFSH (20 ng/ml) or hCG (10 ng/ml) significantly increased CAT enzyme activity; however, neither hCG nor hFSH increased CAT enzyme activity in cells transfected with pSV2-CAT, a reporter plasmid under the control of the SV40 virus promoter and 5' region. The phosphodiesterase inhibitor isobutylmethylxanthine or the nonhydrolyzable cAMP analog cAMP-chlorothiophenyl significantly increased CAT activity in POMC-CAT-transfected granulosa cells. Human FSH stimulated transcription 10, 20, and 40 h after treatment, but FSH stimulation at the two earlier time points was 2.5- to 5.5-fold greater than that at 40 h. Gonadotropin-stimulated steroidogenesis was equivalent in POMC-CAT-transfected granulosa cells, untransfected, and mock-transfected cells. This indicates that transfection left the physiological hormone response intact. These data demonstrate the following. 1) 767 basepairs of the rat POMC gene are enough to confer gonadotropin stimulation on the CAT marker gene in granulosa cells. 2) Although the POMC promotor lacks a well conserved cAMP response element, either of two different pharmacological manipulations of granulosa cells that raise intracellular cAMP can also stimulate POMC-driven CAT expression. 3) Transfected primary cultures of granulosa cells provide a nontransformed, physiologically relevant model with which to study hormone-regulated gene expression.(ABSTRACT TRUNCATED AT 250 WORDS)

  17. 21 CFR 862.1485 - Luteinizing hormone test system.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Luteinizing hormone test system. 862.1485 Section 862.1485 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test...

  18. Lutein and lutein esters in marigold flowers by high performance chromatography

    Institute of Scientific and Technical Information of China (English)

    JIANG Xin-yu; CHEN Long-sheng; ZHOU Chun-shan

    2005-01-01

    Lutein and lutein esters in marigold flowers was quantitatively determined by high performance chromatography (HPLC) with ODS-C18 column. A mixture of CH3CN - CH3OH - CH3COOCH2CH3 with volume ratio of 55: 1: 44 was used as mobile phase at a flow rate of 1.0 mL/min and detection was carried out at 460 nm. The column temperature was about 20 ℃. The contents of lutein and lutein esters were determined by analytical curve of lutein since lutein and lutein esters have the same spectral characteristics. Determination results of hexane extracts and saponified samples of lutein show that the saponification transforms the esters into free lutein. The increase of the content of dipalmitate and palmitate stearate reveals that the reaction includes transesterifications.

  19. Infertility in Female Mice with a Gain-of-Function Mutation in the Luteinizing Hormone Receptor Is Due to Irregular Estrous Cyclicity, Anovulation, Hormonal Alterations, and Polycystic Ovaries.

    Science.gov (United States)

    Hai, Lan; McGee, Stacey R; Rabideau, Amanda C; Paquet, Marilène; Narayan, Prema

    2015-07-01

    The luteinizing hormone receptor, LHCGR, is essential for fertility in males and females, and genetic mutations in the receptor have been identified that result in developmental and reproductive defects. We have previously generated and characterized a mouse model (KiLHR(D582G)) for familial male-limited precocious puberty caused by an activating mutation in the receptor. We demonstrated that the phenotype of the KiLHR(D582G) male mice is an accurate phenocopy of male patients with activating LHCGR mutations. In this study, we observed that unlike women with activating LHCGR mutations who are normal, female KiLHR(D582G) mice are infertile. Mice exhibit irregular estrous cyclicity, anovulation, and precocious puberty. A temporal study from 2-24 wk of age indicated elevated levels of progesterone, androstenedione, testosterone, and estradiol and upregulation of several steroidogenic enzyme genes. Ovaries of KiLHR(D582G) mice exhibited significant pathology with the development of large hemorrhagic cysts as early as 3 wk of age, extensive stromal cell hyperplasia and hypertrophy with luteinization, numerous atretic follicles, and granulosa cell tumors. Ovulation could not be rescued by the addition of exogenous gonadotropins. The body weights of the KiLHR(D582G) mice were higher than wild-type counterparts, but there was no increase in the body fat composition or metabolic abnormalities such as impaired glucose tolerance and insulin resistance. These studies demonstrate that activating LHCGR mutations do not produce the same phenotype in female mice as in humans and clearly illustrate species differences in the expression and regulation of LHCGR in the ovary, but not in the testis. © 2015 by the Society for the Study of Reproduction, Inc.

  20. Radioiodination of chicken luteinizing hormone without affecting receptor binding potency

    Energy Technology Data Exchange (ETDEWEB)

    Kikuchi, M.; Ishii, S. (Waseda Univ., Tokyo (Japan))

    1989-12-01

    By improving the currently used lactoperoxidase method, we were able to obtain radioiodinated chicken luteinizing hormone (LH) that shows high specific binding and low nonspecific binding to a crude plasma membrane fraction of testicular cells of the domestic fowl and the Japanese quail, and to the ovarian granulosa cells of the Japanese quail. The change we made from the original method consisted of (1) using chicken LH for radioiodination that was not only highly purified but also retained a high receptor binding potency; (2) controlling the level of incorporation of radioiodine into chicken LH molecules by employing a short reaction time and low temperature; and (3) fractionating radioiodinated chicken LH further by gel filtration using high-performance liquid chromatography. Specific radioactivity of the final {sup 125}I-labeled chicken LH preparation was 14 microCi/micrograms. When specific binding was 12-16%, nonspecific binding was as low as 2-4% in the gonadal receptors. {sup 125}I-Labeled chicken LH was displaced by chicken LH and ovine LH but not by chicken follicle-stimulating hormone. The equilibrium association constant of quail testicular receptor was 3.6 x 10(9) M-1. We concluded that chicken LH radioiodinated by the present method is useful for studies of avian LH receptors.

  1. Childhood ovarian juvenile granulosa cell tumour

    African Journals Online (AJOL)

    Prof Ezechukwu

    2012-05-12

    May 12, 2012 ... years old of age. We describe a case ... Juvenile granulosa cell tumour a subtype of ovarian stro- mal cell ... A more serious estrogen effects can occur in various end ... usually behave in a benign manner despite having histo-.

  2. Effects of Lutein and Zeaxanthin on LPS-Induced Secretion of IL-8 by Uveal Melanocytes and Relevant Signal Pathways

    OpenAIRE

    Shih-Chun Chao; Tommaso Vagaggini; Chan-Wei Nien; Sheng-Chieh Huang; Hung-Yu Lin

    2015-01-01

    The effects of lutein and zeaxanthin on lipopolysaccharide- (LPS-) induced secretion of IL-8 by uveal melanocytes (UM) were tested in cultured human UM. MTT assay revealed that LPS (0.01–1 μg/mL) and lutein and zeaxanthin (1–10 μM) did not influence the cell viability of cultured UM. LPS caused a dose-dependent increase of secretion of IL-8 by cultured UM. Lutein and zeaxanthin did not affect the constitutive secretion of IL-8. However, lutein and zeaxanthin decreased LPS-induced secretion of...

  3. Production of specific antisera for radioimmunoassay of human luteinizing hormone (LH) in the presence of human chorionic gonadotropin (hCG). [/sup 125/I

    Energy Technology Data Exchange (ETDEWEB)

    Thorell, J.I.; Jeppsson, S.; Holmstrom, B.

    1976-09-01

    A specific radioimmunoassay for LH, which measures plasma LH in the presence of human chorionic gonadotropin (hCG) is described. Rabbits were immunized with highly purified native LH. One of the antisera with a difference in its reactivity against LH and hCG was further purified by affinity chromatography on a column with hCG coupled to Sepharose 4B. The adsorbed antiserum and /sup 125/I-LH was used in a double antibody assay. The LH standard (MRC/68/40) efficiently inhibited the binding of /sup 125/I-LH, and the standard curve showed a sensitivity of 0.5 ng/ml in the sample. hCG up to 10,000 ng/ml did not inhibit the binding of /sup 125/I-LH. The plasma level of LH in pregnant women in the first trimester was low (1.3 +- 0.1 ng/ml). When LH was measured in fertile or menopausal women with or without stimulation with LH/FSH releasing hormone (LH-RH)/sup x/ the results agreed to those found with our conventional LH-assay based on antiserum against hCG.

  4. Premature luteinization during gonadotropin-releasing hormone antagonist cycles and its relationship with in vitro fertilization outcome.

    Science.gov (United States)

    Bosch, Ernesto; Valencia, Iván; Escudero, Ernesto; Crespo, Juana; Simón, Carlos; Remohí, José; Pellicer, Antonio

    2003-12-01

    To determine the prevalence and the effect of premature luteinization in GnRH antagonist IVF-ET cycles. Prospective observational study. In vitro fertilization-embryo transfer (IVF-ET) program at the Instituto Valenciano de Infertilidad. Eighty-one infertile patients undergoing controlled ovarian hyperstimulation with gonadotropins and GnRH antagonist for IVF-ET. Gonadotropin-releasing hormone (GnRH) antagonist was administered from stimulation day 6. Serum P, E(2), and LH were determined on the day of hCG administration. Cycles were grouped according to serum P level on the day of hCG administration ( or =1.2 ng/mL). Clinical pregnancy and implantation rates were determined. The incidence of premature luteinization was 38.3%. Total recombinant FSH dose and stimulation days differed significantly between the groups. Pregnancy rate (25.8% vs. 54.0%) and implantation rate (13.8% vs. 32.0%) were significantly lower in the premature luteinization group. Premature luteinization during GnRH antagonist IVF-ET cycles is a frequent event that is associated with lower pregnancy and implantation rates. Progesterone elevations are not related to serum LH levels and may reflect the mature granulosa cell response to high FSH exposure.

  5. Defective Gonadotropin-Dependent Ovarian Folliculogenesis and Granulosa Cell Gene Expression in Inhibin-Deficient Mice

    Science.gov (United States)

    Nagaraja, Ankur K.; Middlebrook, Brooke S.; Rajanahally, Saneal; Myers, Michelle; Li, Qinglei; Matzuk, Martin M.; Pangas, Stephanie A.

    2010-01-01

    Inhibin-α knockout (Inha−/−) female mice develop sex cord-stromal ovarian cancer with complete penetrance and previous studies demonstrate that the pituitary gonadotropins (FSH and LH) are influential modifiers of granulosa cell tumor development and progression in inhibin-deficient females. Recent studies have demonstrated that Inha−/− ovarian follicles develop precociously to the early antral stage in prepubertal mice without any increase in serum FSH. These studies suggest that in the absence of inhibins, granulosa cells differentiate abnormally and thus at sexual maturity may undergo an abnormal response to gonadotropin signaling contributing to tumor development. To test this hypothesis, we stimulated immature wild-type and Inha−/− female mice with gonadotropin analogs prior to tumor formation and subsequently examined gonadotropin-induced ovarian follicle development as well as preovulatory and human chorionic gonadotropin-induced gene expression changes in granulosa cells. We find that at 3 wk of age, inhibin-deficient ovaries do not show further antral development or undergo cumulus expansion. In addition, there are widespread alterations in the transcriptome of gonadotropin-treated Inha−/− granulosa cells, with significant changes in genes involved in extracellular matrix and cell-cell communication. These data indicate the gonadotropins initiate an improper program of cell differentiation prior to tumor formation in the absence of inhibins. PMID:20739397

  6. Identification of new ovulation-related genes in humans by comparing the transcriptome of granulosa cells before and after ovulation triggering in the same controlled ovarian stimulation cycle

    DEFF Research Database (Denmark)

    Wissing, M L; Kristensen, S G; Andersen, C Y

    2014-01-01

    with ovarian cancer, while down-regulated genes mainly represented cell cycle and proliferation. WHAT IS KNOWN ALREADY: Radical changes occur in the follicle during final follicle maturation after the ovulatory trigger: these range from ensuring an optimal milieu for the oocyte in meiotic arrest to the release...... of a mature oocyte and remodeling into a corpus luteum. A wide range of mediators of final follicle maturation has been identified in rodents, non-human primates and cows. STUDY DESIGN, SIZE, DURATION: Prospective cohort study including 24 women undergoing ovarian stimulation with the long gonadotrophin-releasing...... hormone agonist protocol during 2010-2012 at Holbæk Fertility Clinic. Nine paired samples of GC and 24 paired samples of follicular fluid (FF) were obtained before and after recombinant human chorionic gonadotrophin (rhCG) administration. PARTICIPANTS/MATERIALS, SETTING, METHODS: Nine paired (nine arrays...

  7. Rescuing Cattleya granulosa Lindley in the wild

    OpenAIRE

    Camara-Neto, Clementino; Chaves-Camara, Iuna; Carvalho de Medeiros, Severino; de Almeida Braga, Maria do Rosario

    2015-01-01

    The Orchid Society of Rio Grande do Norte State, Brazil, through its Group of Experimental Interactive Research (SORN/GEPI), has been study- ing the occurrence of Cattleya granulosa Lindley, 1842, specifically in the coastal sand plain and dune vegetation (“restinga”) and in remaining patches of Atlantic Rainforest in the state.  The Orchid Society of Rio Grande do Norte State, Brazil, through its Group of Experimental Interactive Research (SORN/GEPI), has been study- ing the occurrence of...

  8. Differential effect of dietary antioxidant classes (carotenoids, polyphenols, vitamins C and E) on lutein absorption.

    Science.gov (United States)

    Reboul, Emmanuelle; Thap, Sinay; Tourniaire, Franck; André, Marc; Juhel, Christine; Morange, Sophie; Amiot, Marie-Josèphe; Lairon, Denis; Borel, Patrick

    2007-03-01

    Lutein is assumed to protect the human retina from blue light and oxidative stress and diminish the incidence of age-related macular degeneration. This antioxidant is commonly ingested with other dietary antioxidants. The aim of the present study was to assess whether the main dietary antioxidants, i.e. carotenoids, polyphenols and vitamins C and E, affect lutein absorption. We measured the effect of adding a mixture of antioxidants (500 mg vitamin C, 67 mg (100 IU) vitamin E and 1 g polyphenols) to a lutein-containing meal (18 mg) on the postprandial lutein response in the chylomicron-rich fraction in eight healthy men. Lutein response was weakest (-23 %; P=0 x 07) after ingestion of the meal containing antioxidants (21 x 9 (sem 4 x 6) v. 28 x 4 (sem 7 x 2) nmol x h/l). To assess the effect of each class of antioxidants and potential interactions, we subsequently evaluated the effect of various combinations of antioxidants on lutein uptake by human intestinal Caco-2 TC-7 cells. A full factorial design showed that both a mixture of polyphenols (gallic acid, caffeic acid, (+)-catechin and naringenin) and a mixture of carotenoids (lycopene plus beta-carotene) significantly (P<0 x 05) impaired lutein uptake by (-10 to-30 %), while vitamins C and E had no significant effect. Subsequent experiments showed that the aglycone flavanone naringenin was the only polyphenol responsible for the effect of the polyphenol mixture, and that the carotenoid effect was not carotenoid species-dependent. Taken together, the present results suggest that lutein absorption is not markedly affected by physiological concentrations of vitamins C and E but can be impaired by carotenoids and naringenin

  9. Novel protocol for lutein extraction from microalga Chlorella vulgaris

    DEFF Research Database (Denmark)

    D'Este, Martina; De Francisci, Davide; Angelidaki, Irini

    2017-01-01

    Lutein is a pigment generally extracted from marigold flowers. However, lutein is also found in considerable amounts in microalgae. In this study a novel method was developed to improve the extraction efficiency of lutein from microalga C. vulgaris. Differently from conventional methods, ethanol...... was used instead of water in the saponification step, which was conducted simultaneously to the solvent extraction, performed using dichloromethane. The amount of lutein extracted from C. vulgaris dried biomass increased more than threefold, from 0.20 ± 0.00 mgLutein/gDM to 0.69 ± 0.08 mgLutein/gDM. Lutein...

  10. Ultrastructure of the basal lamina of bovine ovarian follicles and its relationship to the membrana granulosa.

    Science.gov (United States)

    Irving-Rodgers, H F; Rodgers, R J

    2000-03-01

    Different morphological phenotypes of follicular basal lamina and of membrana granulosa have been observed. Ten preantral follicles (membrana granulosa. Within each antral follicle, the shape of the basal cells of the membrana granulosa was uniform, and either rounded or columnar. There were equal proportions of follicles membrana granulosa.

  11. Acute and subacute toxicity assessment of lutein in lutein-deficient mice.

    Science.gov (United States)

    Nidhi, Bhatiwada; Baskaran, Vallikannan

    2013-10-01

    Dietary lutein consumption is lower than the actual recommended allowances to prevent macular degeneration; thus dietary lutein supplements have been recommended. This study aimed to investigate potential adverse effect of lutein from Tagetes erecta in lutein-deficient (LD) male mice. Preliminary acute toxicity study revealed that the LD50 exceeded the highest dose of 10000 mg/kg BW. In a subacute study, male mice were gavaged with 0, 100, 1000 mg/kg BW/day for a period of 4 wk. Plasma lutein levels increased dose dependently (P < 0.01) after acute and subacute feeding of lutein in LD mice. Compared to the control (peanut oil without lutein) group, no treatment-related toxicologically significant effects of lutein were prominent in clinical observation, ophthalmic examinations, body, and organ weights. Further, no toxicologically significant findings were eminent in hematological, histopathological, and other clinical chemistry parameters. In the oral subacute toxicity study, the no-observed-adverse-effect level (NOAEL) for lutein in LD mice was determined as 1000 mg/kg/day, the highest dose tested. © 2013 Institute of Food Technologists®

  12. The incidence of apoptotic bodies in membrana granulosa can predict prognosis of ova from patients participating in in vitro fertilization programs.

    Science.gov (United States)

    Nakahara, K; Saito, H; Saito, T; Ito, M; Ohta, N; Takahashi, T; Hiroi, M

    1997-08-01

    To investigate the relationship between the incidence of apoptotic bodies in membrana granulosa and follicular steroid concentrations in human follicles. Case-controlled prospective study for 132 individual follicles. Procedures were performed in Yamagata University Hospital. Thirty-six normo-ovulatory women with tubal infertility underwent ovulation induction for IVF-ET with a conventional hyperstimulation method. Patients underwent follicle aspiration after the administration of hCG. The nuclei of recovered granulosa cells were examined by fluorescence microscopy, and the incidence of apoptotic bodies was tabulated. Intrafollicular steroids were evaluated mainly by RIA. These data were analyzed with respect to oocyte-retrieval, oocyte maturity, fertilization, and embryo quality. Membrana granulosa cells in the follicles from which oocytes were subsequently fertilized showed a significantly lower incidence of apoptotic bodies than those in follicles from which the oocytes did not fertilize. Membrana granulosa cells in the follicles from which oocytes were developed into good quality showed a significantly lower incidence of apoptotic bodies than those in the follicles from which oocytes developed into fair and poor quality. The incidence of apoptotic bodies was significantly higher in the mural granulosa cell region than in the cumulus cell region in most cases. Intrafollicular E2, P, and free T levels were not different between the oocyte groups. These results indicate that lower incidence of apoptotic bodies in individual follicles is associated with better outcomes for oocytes. Also, mural granulosa cells and cumulus cell in each follicle may show differentiation during follicular maturation.

  13. Infertility in Female Mice with a Gain-of-Function Mutation in the Luteinizing Hormone Receptor Is Due to Irregular Estrous Cyclicity, Anovulation, Hormonal Alterations, and Polycystic Ovaries1

    Science.gov (United States)

    Hai, Lan; McGee, Stacey R.; Rabideau, Amanda C.; Paquet, Marilène; Narayan, Prema

    2015-01-01

    The luteinizing hormone receptor, LHCGR, is essential for fertility in males and females, and genetic mutations in the receptor have been identified that result in developmental and reproductive defects. We have previously generated and characterized a mouse model (KiLHRD582G) for familial male-limited precocious puberty caused by an activating mutation in the receptor. We demonstrated that the phenotype of the KiLHRD582G male mice is an accurate phenocopy of male patients with activating LHCGR mutations. In this study, we observed that unlike women with activating LHCGR mutations who are normal, female KiLHRD582G mice are infertile. Mice exhibit irregular estrous cyclicity, anovulation, and precocious puberty. A temporal study from 2–24 wk of age indicated elevated levels of progesterone, androstenedione, testosterone, and estradiol and upregulation of several steroidogenic enzyme genes. Ovaries of KiLHRD582G mice exhibited significant pathology with the development of large hemorrhagic cysts as early as 3 wk of age, extensive stromal cell hyperplasia and hypertrophy with luteinization, numerous atretic follicles, and granulosa cell tumors. Ovulation could not be rescued by the addition of exogenous gonadotropins. The body weights of the KiLHRD582G mice were higher than wild-type counterparts, but there was no increase in the body fat composition or metabolic abnormalities such as impaired glucose tolerance and insulin resistance. These studies demonstrate that activating LHCGR mutations do not produce the same phenotype in female mice as in humans and clearly illustrate species differences in the expression and regulation of LHCGR in the ovary, but not in the testis. PMID:26040673

  14. Ultrasound-Enhanced Subcritical CO2 Extraction of Lutein from Chlorella pyrenoidosa.

    Science.gov (United States)

    Fan, Xiao-Dan; Hou, Yan; Huang, Xing-Xin; Qiu, Tai-Qiu; Jiang, Jian-Guo

    2015-05-13

    Lutein is an important pigment of Chlorella pyrenoidosa with many beneficial functions in human health. The main purpose of this study was to extract lutein from C. pyrenoidosa using ultrasound-enhanced subcritical CO2 extraction (USCCE). Effects of operating conditions on the extraction, including extraction pretreatment, temperature, pressure, time, CO2 flow rate, and ultrasonic power, were investigated, and an orthogonal experiment was designed to study the effects of extraction pressure, temperature, cosolvent amount, and time on the extraction yields. The USCCE method was compared with other extraction methods in terms of the yields of lutein and the microstructure of C. pyrenoidosa powder by scanning electron microscopy. A maximal extraction yield of 124.01 mg lutein/100 g crude material was achieved under optimal conditions of extraction temperature at 27 °C, extraction pressure at 21 MPa, cosolvent amount at 1.5 mL/g ethanol, and ultrasound power at 1000 W. Compared to other methods, USCCE could significantly increase the lutein extraction yield at lower extraction temperature and pressure. Furthermore, the kinetic models of USCCE and subcritical CO2 extraction (SCCE) of lutein from C. pyrenoidosa were set as E = 130.64 × (1 - e(-0.6599t)) and E = 101.82 × (1 - e(-0.5683t)), respectively. The differences of parameters in the kinetic models indicate that ultrasound was able to enhance the extraction process of SCCE.

  15. Olive oil improves the intestinal absorption and bioavailability of lutein in lutein-deficient mice.

    Science.gov (United States)

    Nidhi, Bhatiwada; Mamatha, Bangera Sheshappa; Baskaran, V

    2014-02-01

    To investigate the influence of olive (OO), groundnut (GNO), soybean (SBO), sunflower (SFO), rice bran (RBO), corn (CO), palm (PO) oil or mixed micelle (control) on absorption kinetics and bioavailability of lutein in lutein-deficient mice. Additional aim was to correlate the activity of intestinal triacylglycerol lipase with intestinal and plasma lutein levels. After induction of lutein deficiency, mice (n = 165) were divided into eight groups (OO, SFO, GNO, RBO, PO, CO, SBO and control; n = 20/group) and the remaining (n = 5) were used as baseline (0 h). Groups were further divided into four subgroups (n = 5/subgroup) and were intubated with lutein (200 μM) dispersed in different vegetable oils. Plasma and tissue (intestine, liver and eyes), lutein, triglycerides, intestinal triacylglycerol lipases and fatty acid profile of plasma and tissues were measured at different time intervals. The percentage area under the curve value for plasma lutein in OO and GNO was higher by 41.8 and 5.1 %, while it was lower in other groups (18.2-53.3 %), when compared to control. Similarly, the percentage area under the curve for eye lutein in OO and GNO groups was higher by 35.2 and 4.8 %, whereas in other groups it was lower (5.4-69 %) than in control. Results show that olive oil facilitates the lutein absorption more compared to other vegetable oils, which may be due to the difference in fatty acid composition and higher activity of intestinal triacylglycerol lipase. Dietary olive oil rich in oleic acid improves the bioavailability and accumulation of lutein in lutein-deficient mice by modifying the intestinal triacylglycerol lipase activity.

  16. Luteinizing Hormone Receptor Knockout (LuRKO) Mice and Transgenic Human Chorionic Gonadotropin (hCG)-Overexpressing Mice (hCG αβ+) Have Bone Phenotypes

    National Research Council Canada - National Science Library

    Yarram, S. J; Perry, M. J; Christopher, T. J; Westby, K; Brown, N. L; Lamminen, T; Rulli, S. B; Zhang, F.-P; Huhtaniemi, I; Sandy, J. R; Mansell, J. P

    2003-01-01

    Considerable attention has been paid to the role of sex steroids during periods of major skeletal turnover, but the interaction of the gonadotropic hormones, which include LH, FSH, and human chorionic gonadotropin (hCG...

  17. Ovarian reserve status in young women is associated with altered gene expression in membrana granulosa cells.

    Science.gov (United States)

    Skiadas, Christine C; Duan, Shenghua; Correll, Mick; Rubio, Renee; Karaca, Nilay; Ginsburg, Elizabeth S; Quackenbush, John; Racowsky, Catherine

    2012-07-01

    Diminished ovarian reserve (DOR) is a challenging diagnosis of infertility, as there are currently no tests to predict who may become affected with this condition, or at what age. We designed the present study to compare the gene expression profile of membrana granulosa cells from young women affected with DOR with those from egg donors of similar age and to determine if distinct genetic patterns could be identified to provide insight into the etiology of DOR. Young women with DOR were identified based on FSH level in conjunction with poor follicular development during an IVF cycle (n = 13). Egg donors with normal ovarian reserve (NOR) comprised the control group (n = 13). Granulosa cells were collected following retrieval, RNA was extracted and microarray analysis was conducted to evaluate genetic differences between the groups. Confirmatory studies were undertaken with quantitative RT-PCR (qRT-PCR). Multiple significant differences in gene expression were observed between the DOR patients and egg donors. Two genes linked with ovarian function, anti-Mullerian hormone (AMH) and luteinizing hormone receptor (LHCGR), were further analyzed with qRT-PCR in all patients. The average expression of AMH was significantly higher in egg donors (adjusted P-value = 0.01), and the average expression of LHCGR was significantly higher in DOR patients (adjusted P-value = 0.005). Expression levels for four additional genes, progesterone receptor membrane component 2 (PGRMC2), prostaglandin E receptor 3 (subtype EP3) (PTGER3), steroidogenic acute regulatory protein (StAR), and StAR-related lipid transfer domain containing 4 (StarD4), were validated in a group consisting of five NOR and five DOR patients. We conclude that gene expression analysis has substantial potential to determine which young women may be affected with DOR. More importantly, our analysis suggests that DOR patients fall into two distinct subgroups based on gene expression profiles, indicating that different

  18. Down-regulation of membrana granulosa cell gap junctions is correlated with irreversible commitment to resume meiosis in golden Syrian hamster oocytes.

    Science.gov (United States)

    Racowsky, C; Baldwin, K V; Larabell, C A; DeMarais, A A; Kazilek, C J

    1989-08-01

    One of the currently popular hypotheses for the regulation of meiotic resumption in mammalian oocytes proposes that the preovulatory surge of luteinizing hormone causes down-regulation of follicular gap junctions, which in turn disrupts transfer of a meiotic arrester from the somatic cells into the oocyte. The present study has investigated this hypothesis by examining the integrity of membrana granulosa cell gap junctions during the period of irreversible commitment to maturation of golden Syrian hamster oocytes in vivo. Our results have revealed a significant progressive decrease in the fractional area of cell surface occupied by gap junction membrane with increasing percentage of oocytes irreversibly committed to mature (1.946% and 0.921% fractional gap junction area at 0% and 100% oocytes irreversibly committed to mature, respectively, P less than 0.05). This net loss of membrana granulosa cell gap junctions from the cell surface was accompanied by a significant decrease in density of gap junction particles, whether they were arranged in rectilinear or non-rectilinear packing patterns. Furthermore, the number of gap junction particles per unit area of surface membrane scanned also underwent a significant progressive decrease with increasing percentage of oocytes irreversibly committed to mature. These data with the hamster are consistent with the hypothesis that down-regulation of membrana granulosa cell gap junctions may be of central importance in the regulation of gonadotropic stimulation of meiotic resumption in mammalian oocytes.

  19. Effect of epidermal growth factor on follicle-stimulating hormone-induced proliferation of granulosa cells from chicken prehierarchical follicles

    Institute of Scientific and Technical Information of China (English)

    Jin-xing LIN; Yu-dong JIA; Cai-qiao ZHANG

    2011-01-01

    The development of ovarian follicular cells is controlled by multiple circulating and local hormones and factors,including follicle-stimulating hormone (FSH) and epidermal growth factor (EGF).In this study,the stagespecific effect of EGF on FSH-induced proliferation of granulosa cells was evaluated in the ovarian follicles of egg-laying chickens.Results showed that EGF and its receptor (EGFR) mRNAs displayed a high expression in granulosa cells from the prehierarchical follicles,including the large white follicle (LWF) and small yellow follicle (SYF),and thereafter the expression decreased markedly to the stage of the largest preovulatory follicle.SYF represents a turning point of EGF/EGFR mRNA expression during follicle selection.Subsequently the granulosa cells from SYF were cultured to reveal the mediation of EGF in FSH action.Cell proliferation was remarkably increased by treatment with either EGF or FSH (0.1-100 ng/ml).This result was confirmed by elevated proliferating cell nuclear antigen (PCNA) expression and decreased cell apoptosis.Furthermore,EGF-induced cell proliferation was accompanied by increased mRNA expressions of EGFR,FSH receptor,and the cell cycle-regulating genes (cyclins D1 and E1,cyclin-dependent kinases 2 and 6) as well as decreased expression of luteinizing hormone receptor mRNA.However,the EGF or FSH-elicited effect was reversed by simultaneous treatment with an EGFR inhibitor AG1478.In conclusion,EGF and EGFR expressions manifested stage-specific changes during follicular development and EGF mediated FSH-induced cell proliferation and retarded cell differentiation in the prehierarchical follicles.These expressions thus stimulated follicular growth before selection in the egg-laying chicken.

  20. Macular lutein and zeaxanthin are related to brain lutein and zeaxanthin in primates

    Science.gov (United States)

    The xanthophyll pigments lutein and zeaxanthin cross the blood-retina barrier to preferentially accumulate in the macular region of the neural retina. There they form macular pigment, protecting the retina from blue light damage and oxidative stress. Lutein and zeaxanthin also accumulate in brain t...

  1. Lutein production from biomass: marigold flowers versus microalgae.

    Science.gov (United States)

    Lin, Jian-Hao; Lee, Duu-Jong; Chang, Jo-Shu

    2015-05-01

    Microalgae have faster growth rates and more free lutein than marigold flowers, the current source of lutein. However, no commercial lutein production uses microalgae. This review compares lutein content, cultivation, harvesting, cell disruption, and extraction stages of lutein production using marigold flowers and those using microalgae as feedstock. The lutein production rate of microalgae is 3-6 times higher than that of marigold flowers. To produce 1 kg of pure lutein, marigolds need more land and water, but require less nutrients (N, P, K) and less energy than microalgae. Since lutein is tightly bound in microalgae and microalgae are small, cell disruption and subsequent extraction stages consume a considerable amount of energy. Research and development of affordable lutein production from microalgae are discussed.

  2. Delayed menopause due to granulosa cell tumor of the ovary

    Directory of Open Access Journals (Sweden)

    Bhushan Murkey

    2011-01-01

    Full Text Available A 52-year-old patient presented with complaints of menorrhagia. Endometrial biopsy revealed simple hyperplasia of the endometrium. Total abdominal hysterectomy with bilateral oophorectomy was carried out. The ovaries looked grossly normal, but histopathology reported granulosa cell tumor of the right ovary. Granulosa cell tumors belong to the sexcord stromal category and account for approximately 2% of all ovarian tumors. We review the features and treatment of granulosa cell tumors and the importance of screening for ovarian tumors in a case of endometrial hyperplasia and delayed menopause.

  3. Zearalenone exposure affects mouse oocyte meiotic maturation and granulosa cell proliferation.

    Science.gov (United States)

    Hou, Yan-Jun; Zhu, Cheng-Cheng; Xu, Yin-Xue; Cui, Xiang-Shun; Kim, Nam-Hyung; Sun, Shao-Chen

    2015-09-01

    Zearalenone (ZEN) is a metabolite of Fusarium and is a common contaminant of grains and foodstuffs. ZEN acts as a xenoestrogen and is considered to be cytotoxic, tissue toxic, and genotoxic, which causes abortions and stillbirths in humans and animals. Since estrogens affect oocyte maturation during meiosis, in this study we investigated the effects of ZEN on mouse oocyte meiotic maturation and granulosa cell proliferation. Our results showed that ZEN-treated oocyte maturation rates were decreased, which might be due to the disrupted cytoskeletons: (1) ZEN treatment resulted in significantly more oocytes with abnormal spindle morphologies; (2) actin filament expression and distribution were also disrupted after ZEN treatment, which was confirmed by the aberrant distribution of actin regulatory proteins. In addition, cortical granule-free domains (CGFDs) were disrupted after ZEN treatment, which indicated that ZEN may affect mouse oocyte fertilization capability. ZEN reduced mouse granulosa cell proliferation in a dose-dependent manner as determined by MTT assay and TUNEL apoptosis analysis, which may be another cause for the decreased oocyte maturation. Thus, our results demonstrated that exposure to zearalenone affected oocyte meiotic maturation and granulosa cell proliferation in mouse. © 2014 Wiley Periodicals, Inc.

  4. Generation of iPS Cells from Granulosa Cells.

    Science.gov (United States)

    Mao, Jian; Liu, Lin

    2016-01-01

    Various types of somatic cells can be reprogrammed to induced pluripotent stem (iPS) cells. Somatic stem cells may generate iPS cells more efficiently than do differentiated cells. We show that granulosa cells exhibit characteristic of somatic stem cells and can be reprogrammed to iPS cells more efficiently or with few factors. Here, we describe generation of mouse and pig iPS cells from granulosa cells with high efficiency.

  5. Dietary Sources of Lutein and Zeaxanthin Carotenoids and Their Role in Eye Health

    Directory of Open Access Journals (Sweden)

    Rashida Ali

    2013-04-01

    Full Text Available The eye is a major sensory organ that requires special care for a healthy and productive lifestyle. Numerous studies have identified lutein and zeaxanthin to be essential components for eye health. Lutein and zeaxanthin are carotenoid pigments that impart yellow or orange color to various common foods such as cantaloupe, pasta, corn, carrots, orange/yellow peppers, fish, salmon and eggs. Their role in human health, in particular the health of the eye, is well established from epidemiological, clinical and interventional studies. They constitute the main pigments found in the yellow spot of the human retina which protect the macula from damage by blue light, improve visual acuity and scavenge harmful reactive oxygen species. They have also been linked with reduced risk of age-related macular degeneration (AMD and cataracts. Research over the past decade has focused on the development of carotenoid-rich foods to boost their intake especially in the elderly population. The aim of this article is to review recent scientific evidences supporting the benefits of lutein and zexanthin in preventing the onset of two major age-related eye diseases with diets rich in these carotenoids. The review also lists major dietary sources of lutein and zeaxanthin and refers to newly developed foods, daily intake, bioavailability and physiological effects in relation to eye health. Examples of the newly developed high-lutein functional foods are also underlined.

  6. Concanavalin-A induces granulosa cell death and inhibits FSH-mediated follicular growth and ovarian maturation in female rats.

    Science.gov (United States)

    Velasquez, Ethel V; Ríos, Mariana; Ortiz, María Elena; Lizama, Carlos; Nuñez, Elizabeth; Abramovich, Dalhia; Orge, Felipe; Oliva, Barbara; Orellana, Renán; Villalon, Manuel; Moreno, Ricardo D; Tesone, Marta; Rokka, Anne; Corthals, Garry; Croxatto, Horacio B; Parborell, Fernanda; Owen, Gareth I

    2013-05-01

    Reproductive success stems from a finely regulated balance between follicular maturation and atresia, in which the role of carbohydrate structure is poorly understood. Here, we describe for the first time a fraction of purified recombinant human FSH that is capable of bringing about the cell death of granulosa cells and preventing follicular maturation in a rat model. Further analysis by mass spectrometry revealed the presence of the lectin Concanavalin-A (Con-A) within this fraction of recombinant FSH. Using both the fractionated FSH and Con-A, the observed cell death was predominantly located to the granulosa cells. Ex vivo culture of rat follicles demonstrated that follicle degeneration occurred and resulted in the release of a denuded and deteriorated oocyte. Moreover, in vivo experiments confirmed an increase in atresia and a corresponding reduction confined to follicle in early antral stage. As a mechanism of action, Con-A reduces ovarian proliferation, Von Willebrand staining, and angiogenesis. Based on the observation that Con-A may induce granulosa cell death followed by follicle death, our results further demonstrate that follicular carbohydrate moiety is changing under the influence of FSH, which may allow a carbohydrate-binding lectin to increase granulosa cell death. The physiological consequences of circulating lectin-like molecules remain to be determined. However, our results suggest a potential exploitation of carbohydrate binding in fertility and ovarian cancer treatment. This work may shed light on a key role of carbohydrates in the still obscure physiological process of follicular selection and atresia.

  7. [Precocious pseudopuberty secondary to granulosa cell tumor].

    Science.gov (United States)

    Fernández, F; Jordán, J; Carmona, M; Oliver, A; Gracia, R; González, M; Peralta, A

    1984-12-01

    A case report of pseudoprecocity secondary to a unilateral ovarian tumor of granulosa cells is presented in a 13 month old female. Clinical manifestations appeared at two months of age as unilateral enlargement of the breast, development of pubic hair and vaginal discharge. Plasma estrogen levels were elevated, whereas there was no response of FSH and LH to LH-RH stimulation. The absence of a palpable abdominal mass and a normal ultrasound examination of the abdomen must be pointed out in our case. The suspected clinical and laboratory diagnosis was later confirmed by surgical abdominal examination and ovarian histopathology study. With the exception of a minimal breast enlargement which persists at two years of age, all other signs of pseudoprecocity have disappeared after the surgical removal of the neoplasm. The importance of surgical abdominal examination must be pointed out as a diagnostic method when clinical and laboratory findings suggest an ovarian tumor inspite of normal abdominal palpation, ultrasound and roentgenology.

  8. Effects of Progestin and Antiprogestin on the Expression of FSH Receptor and LH Receptor mRNA in Porcine Granulosa and Thecal Cells

    Institute of Scientific and Technical Information of China (English)

    吴尔若; 刘德瑜; 赵金来; 吴燕婉

    2000-01-01

    In order to investigate the mechanism of progestin and antiprogestin in the regula-tion of ovarian steroidogenesis, a dual-chamber culture system was prepared with the amnion membrane of human placenta. Isolated porcine granulosa and thecal cells from 4~6 mm-diameter follicles were grown on both sides of the amnion, respectively, and co-cultured with or without LNG and RU486. After 48 h incubation, the mRNAs of FSH receptor (FSH-R) and LH receptor (LH-R) of both cells were observed by in situ hybridization. The results showed that granulosa cells expressed both FSH-R mR-NA and LH-R mRNA, while thecal cells expressed LH-R mRNA only. Under the stimulation of FSH, both LNG and RU486 increased FSH-R mRNA expression of granulosa cells. Under the stimulation of LH, LNG enhanced LH-R mRNA expres-sion of thecal cells;while RU486 decreased its expression. When granulosa and thecal cells were exposed to FSH and LH both, the actions of LNG and RU 486 in thecal cells showed the same result as that stimulated by LH alone. In granulosa cells LNG de-creased LH-R mRNA expression, while RU486 increased its expression. These data suggest that; (1) granulosa cells expressed FSH-R mRNA significantly; (2) both the progestin and antiprogestin directly acted on the mRNA expression of gonadotropin re-ceptors of ovarian cells, but effects were different; (3) the response of granulosa or thecal cells to the action of LNG and RU486 was not the same. The mechanism needs to be further investigated.

  9. Effects of Lutein and Zeaxanthin on LPS-Induced Secretion of IL-8 by Uveal Melanocytes and Relevant Signal Pathways

    Directory of Open Access Journals (Sweden)

    Shih-Chun Chao

    2015-01-01

    Full Text Available The effects of lutein and zeaxanthin on lipopolysaccharide- (LPS- induced secretion of IL-8 by uveal melanocytes (UM were tested in cultured human UM. MTT assay revealed that LPS (0.01–1 μg/mL and lutein and zeaxanthin (1–10 μM did not influence the cell viability of cultured UM. LPS caused a dose-dependent increase of secretion of IL-8 by cultured UM. Lutein and zeaxanthin did not affect the constitutive secretion of IL-8. However, lutein and zeaxanthin decreased LPS-induced secretion of IL-8 in cultured UM in a dose-dependent manner. LPS significantly increased NF-κB levels in cell nuclear extracts and p-JNK levels in the cell lysates from UM, but not p-p38 MAPK and p-ERG. Lutein or zeaxanthin significantly reduced LPS-induced increase of NF-κB and p-JNK levels, but not p38 MAPK and ERG levels. The present study demonstrated that lutein and zeaxanthin inhibited LPS-induced secretion of IL-8 in cultured UM via JNK and NF-κB signal pathways. The anti-inflammatory effects of lutein and zeaxanthin might be explored as a therapeutic approach in the management of uveitis and other inflammatory diseases of the eye.

  10. Effects of Lutein and Zeaxanthin on LPS-Induced Secretion of IL-8 by Uveal Melanocytes and Relevant Signal Pathways.

    Science.gov (United States)

    Chao, Shih-Chun; Vagaggini, Tommaso; Nien, Chan-Wei; Huang, Sheng-Chieh; Lin, Hung-Yu

    2015-01-01

    The effects of lutein and zeaxanthin on lipopolysaccharide- (LPS-) induced secretion of IL-8 by uveal melanocytes (UM) were tested in cultured human UM. MTT assay revealed that LPS (0.01-1 μg/mL) and lutein and zeaxanthin (1-10 μM) did not influence the cell viability of cultured UM. LPS caused a dose-dependent increase of secretion of IL-8 by cultured UM. Lutein and zeaxanthin did not affect the constitutive secretion of IL-8. However, lutein and zeaxanthin decreased LPS-induced secretion of IL-8 in cultured UM in a dose-dependent manner. LPS significantly increased NF-κB levels in cell nuclear extracts and p-JNK levels in the cell lysates from UM, but not p-p38 MAPK and p-ERG. Lutein or zeaxanthin significantly reduced LPS-induced increase of NF-κB and p-JNK levels, but not p38 MAPK and ERG levels. The present study demonstrated that lutein and zeaxanthin inhibited LPS-induced secretion of IL-8 in cultured UM via JNK and NF-κB signal pathways. The anti-inflammatory effects of lutein and zeaxanthin might be explored as a therapeutic approach in the management of uveitis and other inflammatory diseases of the eye.

  11. Significance of lutein in red blood cells of Alzheimer's disease patients.

    Science.gov (United States)

    Kiko, Takehiro; Nakagawa, Kiyotaka; Tsuduki, Tsuyoshi; Suzuki, Toshihide; Arai, Hiroyuki; Miyazawa, Teruo

    2012-01-01

    Red blood cells (RBC) of Alzheimer's disease (AD) patients are known to be in an excessively oxidized state (i.e., with a high accumulation of peroxidized phospholipids (PLOOH)). Previously we confirmed in vitro, in vivo murine, and in human studies that carotenoids can effectively inhibit accumulation of RBC PLOOH. Thus, the relationship between RBC carotenoids and PLOOH concentrations in AD patients is of interest. In this study, RBC carotenoids and PLOOH were evaluated in 28 normal control subjects (age: 74.1 ± 1.3 years) and 28 patients with AD (age: 72.5 ± 1.4 years). The concentrations of RBC carotenoids, especially lutein, in AD patients were significantly lower than in control subjects. An inverse relationship was seen between RBC carotenoids, especially lutein, and PLOOH concentrations in AD patients. These results suggest that RBC lutein, in particular, may contribute to suppression of PLOOH accumulation in RBC of AD patients.

  12. Effect of dietary α-tocopherol on the bioavailability of lutein in laying hen.

    Science.gov (United States)

    Islam, K M S; Khalil, M; Männer, K; Raila, J; Rawel, H; Zentek, J; Schweigert, F J

    2016-10-01

    Lutein and its isomer zeaxanthin have gained considerable interest as possible nutritional ingredient in the prevention of age-related macular degeneration (AMD) in humans. Egg yolk is a rich source of these carotenoids. As an oxidative sensitive component, antioxidants such as α-tocopherol (T) might contribute to an improved accumulation in egg yolk. To test this, chickens were fed lutein esters (LE) with and without α-tocopherol as an antioxidant. After depletion on a wheat-soya bean-based lutein-poor diet for 21 days, laying hens (n = 42) were equally divided into three groups and fed the following diets for 21 days: control (basal diet), a LE group (40 mg LE/kg feed) and LE + T group (40 mg LE plus 100 mg T/kg feed). Eggs and blood were collected periodically. Carotenoids and α-tocopherol in yolk and blood plasma were determined by HPLC. Egg yolk was also analysed for total carotenoids using a one-step spectrophotometric method (iCheck((™)) ). Lutein, zeaxanthin, α-tocopherol and total carotenoids in egg yolk were highest after 14 days of feeding and decreased slightly afterwards. At the end of the trial, eggs of LE + T group contained higher amount of lutein (13.72), zeaxanthin (0.65), α-tocopherol (297.40) and total carotenoids (21.6) compared to the LE group (10.96, 0.55, 205.20 and 18.0 mg/kg, respectively, p < 0.05). Blood plasma values of LE + T group contain higher lutein (1.3), zeaxanthin (0.06) and tocopherol (20.1) compared to LE group (1.02, 0.04 and 14.90 mg/l, respectively, p < 0.05). In conclusion, dietary α-tocopherol enhances bioavailability of lutein reflecting higher content in egg yolk and blood plasma. Improved bioavailability might be due to increased absorption of lutein in the presence of tocopherol and/or a greater stability of lutein/zeaxanthin due to the presence of α-tocopherol as an antioxidant.

  13. The mare as a model for luteinized unruptured follicle syndrome : intrafollicular endocrine milieu

    NARCIS (Netherlands)

    Bashir, S T; Gastal, M O; Tazawa, S P; Tarso, S G S; Hales, D B; Cuervo-Arango, J; Baerwald, A R; Gastal, E L

    2016-01-01

    Luteinized unruptured follicle (LUF) syndrome is a recurrent anovulatory dysfunction that affects up to 23% of women with normal menstrual cycles and up to 73% with endometriosis. Mechanisms underlying the development of LUF syndrome in mares were studied to provide a potential model for human

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  16. Extraordinarily Prolonged Disease Recurrence in a Granulosa Cell Tumor Patient

    Directory of Open Access Journals (Sweden)

    Lisa N. Abaid

    2010-09-01

    Full Text Available Background: Granulosa cell tumors are rare sex cord stromal lesions that comprise approximately 3% of all ovarian neoplasms. The vast majority of granulosa cell tumors are considered indolent but in spite of aggressive management, delayed recurrence is of significant concern. Case Report: We describe a case involving a 67-year-old woman who presented with abdominal pain, bloody stools, and mild nausea. Following a CT scan of the abdomen and pelvis, a 19-cm pelvic mass was identified. Her prior medical history included a hysterectomy for uterine fibroids 40 years ago and a bilateral salpingo-oophorectomy for a presumed granulosa cell tumor 20 years ago. Final pathology revealed granulosa cell tumor with small bowel mesentery involvement. The patient underwent surgical resection and adjuvant chemotherapy; she is currently doing well. Conclusion: Granulosa cell tumors are considered to be of low malignant potential but they have the capacity to recur, even several years following initial patient management. This case exemplifies the disease’s capacity for prolonged recurrence and further accentuates the significance of long-term follow-up in these patients.

  17. Extracellular matrix of the bovine ovarian membrana granulosa.

    Science.gov (United States)

    Rodgers, R J; Irving Rodgers, H F

    2002-05-31

    Much is known about the control of the development of ovarian follicles by growth factors and hormones. The study of extracellular matrix in the ovary, though, is a relatively new area. To date much research has focused on identifying the matrix components present, and more recently, its production and the physiological roles. In this review we focus on the changes that occur in the follicular basal lamina from primordial follicles through to ovulation and formation of the corpus luteum, the changes that occur during follicular atresia, and we discuss our observations of a novel matrix which forms in the membrana granulosa. The follicular basal lamina changes considerably during follicular development in its expression pattern of type IV collagens. Of the laminin chains examined, there appears only to be an increase in amount, except for laminin alpha2. It is expressed only in a small proportion of healthy antral follicles and in the majority of atretic antral follicles. Call-Exner bodies have the same composition as the basal lamina, except they do not contain laminin alpha2, even when the follicular basal lamina does. The novel matrix that develops within the membrana granulosa is similar in composition to Call-Exner bodies which occur predominantly in preantral follicles, except that it is far more common in large antral follicles, does not induce polarization of the surrounding granulosa cells, and does not contain follicular fluid-like material as the Call-Exner bodies of some species do. The expression of this matrix occurs prior to and during the time when granulosa cells express steroidogenic enzymes. It does not exist in corpora lutea. In addition large luteal cells, derived from granulosa cells, do not appear to have a basal lamina. These findings suggest that the maturational changes in the membrana granulosa are accompanied by changes in the matrix.

  18. Lutein: more than just a filter for blue light.

    Science.gov (United States)

    Kijlstra, Aize; Tian, Yuan; Kelly, Elton R; Berendschot, Tos T J M

    2012-07-01

    Lutein is concentrated in the primate retina, where together with zeaxanthin it forms the macular pigment. Traditionally lutein is characterized by its blue light filtering and anti-oxidant properties. Eliminating lutein from the diet of experimental animals results in early degenerative signs in the retina while patients with an acquired condition of macular pigment loss (Macular Telangiectasia) show serious visual handicap indicating the importance of macular pigment. Whether lutein intake reduces the risk of age related macular degeneration (AMD) or cataract formation is currently a strong matter of debate and abundant research is carried out to unravel the biological properties of the lutein molecule. SR-B1 has recently been identified as a lutein binding protein in the retina and this same receptor plays a role in the selective uptake in the gut. In the blood lutein is transported via high-density lipoproteins (HDL). Genes controlling SR-B1 and HDL levels predispose to AMD which supports the involvement of cholesterol/lutein transport pathways. Apart from beneficial effects of lutein intake on various visual function tests, recent findings show that lutein can affect immune responses and inflammation. Lutein diminishes the expression of various ocular inflammation models including endotoxin induced uveitis, laser induced choroidal neovascularization, streptozotocin induced diabetes and experimental retinal ischemia and reperfusion. In vitro studies show that lutein suppresses NF kappa-B activation as well as the expression of iNOS and COX-2. Since AMD has features of a chronic low-grade systemic inflammatory response, attention to the exact role of lutein in this disease has shifted from a local effect in the eye towards a possible systemic anti-inflammatory function.

  19. In vitro and in vivo studies reveal that hamster oocyte meiotic arrest is maintained only transiently by follicular fluid, but persistently by membrana/cumulus granulosa cell contact.

    Science.gov (United States)

    Racowsky, C; Baldwin, K V

    1989-08-01

    Studies were carried out with the golden Syrian hamster to investigate the capacity of follicular fluid to maintain oocyte meiotic arrest and to determine the importance of cumulus-membrana granulosa cell contact in the regulation of meiotic status. The follicular fluid studies were conducted by cytological assessment of meiotic stage up to 6 hr after transferring cumulus-free oocytes into antra of explanted "host" follicles in vitro or into follicles of anesthetized animals prior to the gonadotropin surge at proestrus in vivo. The cumulus-membrana granulosa contact studies were undertaken with explanted follicles in which the oocyte-cumulus complex was dislodged from the underlying membrana granulosa, released into the antrum, and subsequently allowed to reestablish contact during 6 hr of incubation within the follicle. The extent of recontact of the dislodged complex with the underlying membrana granulosa was assessed visually at the end of incubation and was classified as close, moderate, or none. These various degrees of contact typically involved the following number of cumulus cells, as determined by serial sectioning of a representative sample of follicles after dislodgement and subsequent incubation: close, 32.7 +/- 1.78; moderate, 9.0 +/- 2.1; and no contact, 0. After 6 hr of incubation either in vitro or in vivo, few transferred oocytes remained at the germinal vesicle (GV) stage (18.8 +/- 8.7 and 17.3 +/- 4.0% GV, respectively). However, time course experiments revealed that meiotic resumption was significantly delayed in transferred oocytes compared with either liberated oocytes, spontaneously maturing oocytes, or follicle-enclosed oocytes induced to mature by luteinizing hormone in vitro (after 4 hr, transferred, 31.3 +/- 6.0% GV; liberated, 0% GV; follicle-enclosed, 0% GV; after 6 hr, 0% transferred oocytes exhibited a GV). In the dislodgement studies, after 6 hr of incubation, 26% of complexes reestablished close contact with the underlying membrana

  20. Granulosa cell tumor of scrotal tunics: a case report

    Energy Technology Data Exchange (ETDEWEB)

    Ji, Eun Kyung; Cho, Kyoung Sik [Pochon CHA University, Seoul (Korea, Republic of)

    2001-06-01

    We report a case of adult granulosa cell tumor arising in the scrotal tunics. The patient was a 34-year-old man who presented with right scrotal swelling, first noticed four months previously. Under the initial clinical impression of epididymoorchitis, antibiotic treatment was instituted but there was no response. The paratesticular nodules revealed by ultrasound and magnetic resonance imaging mimicked intratesticular lesion, and radical orchiectomy was performed. Although several cases of adult testicular granulosa cell tumor, have been reported, the occurrence of this entity in the paratesticular area has not, as far as we are aware, been previously described.

  1. Highly potent metallopeptide analogues of luteinizing hormone-releasing hormone

    Energy Technology Data Exchange (ETDEWEB)

    Bajusz, S.; Janaky, T.; Csernus, V.J.; Bokser, L.; Fekete, M.; Srkalovic, G.; Redding, T.W.; Schally, A.V. (Tulane Univ. School of Medicine, New Orleans, LA (USA))

    1989-08-01

    Metal complexes related to the cytotoxic complexes cisplatin (cis-diamminedichloroplatinum(II)) and transbis(salicylaldoximato)copper(II) were incorporated into suitably modified luteinizing hormone-releasing hormone (LH-RH) analogues containing D-lysine at position 6. Some of the metallopeptides thus obtained proved to be highly active LH-RH agonists or antagonists. Most metallopeptide analogues of LH-RH showed high affinities for the membrane receptors of rat pituitary and human breast cancer cells. Some of these metallopeptides had cytotoxic activity against human breast cancer and prostate cancer and prostate cancer cell lines in vitro. Such cytostatic metallopeptides could be envisioned as targeted chemotherapeutic agents in cancers that contain receptors for LH-RH-like peptides.

  2. MDR1 overexpression inhibits chemotherapy-induced toxicity of granulosa cells

    Science.gov (United States)

    Salih, Sana M

    2011-01-01

    OBJECTIVE To protect granulosa cells from chemotherapy-induced toxicity by retrovirus-mediated multidrug resistance gene (MDR1) transfection. DESIGN Laboratory study. SETTING Academic research laboratory in a university hospital. INTERVENTION(S) KK15 immortalized murine granulosa cell line was transiently transduced with sf91m3 retrovirus vector carrying MDR1 cDNA that encodes P-glycoprtoein (P-gp). Transduced cells were selected with colchicine and treated with doxorubicin or paclitaxel for 24–72 hours. The expression and function of MDR1 and the mRNA expression of selected steroidogenesis enzymes were evaluated by flow cytometry, cell viability assays, Western blot, and RT-PCR. MAIN OUTCOME MEASURE(S) Viability of sf91m3-transduced KK15 cells after treatment with doxorubicin and paclitaxel. RESULT(S) sf91m3-transduced KK15 demonstrated high expression of biologically active MDR1 as shown by flow cytometry analysis and immunoblotting using P-gp monoclonal antibody and Rhodamine 123 efflux assays. sf91m3-transduced KK15 exhibited significant resistance to toxicity of 10uM paclitaxel(p≤0.001). MDR1-transduced KK15 cells were also protected from doxorubicin toxicity (10nM to 2.5uM) as shown by cell viability assay (p≤0.02). Both flow cytometry and cell viability assay showed that the protection of KK15 from doxorubicin toxicity was lost at 5 uM of doxorubicin; equivalent to 500 times LD50 (p≥0.05). sf91m3-transduced KK15 showed normal mRNA expression of a panel of selected steroidogenesis enzymes. CONCLUSION(S) Retroviral gene delivery of human MDR1 inhibited chemotherapy- induced granulosa cell toxicity and offered chemoprotection in an in vitro model. PMID:21316663

  3. DOSE-RESPONSE OF PORCINE OVARIAN GRANULOSA CELLS TO AMYGDALIN TREATMENT COMBINED WITH DEOXYNIVALENOL

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    Marek Halenár

    2014-02-01

    Full Text Available Amygdalin is one of many nitrilosides, which are natural cyanide-containing substances abundant in the seeds of apricots, almond, peaches, apples, and other rosaceous plants. It is a controversial anti-tumor natural product that has been used as an alternative cancer drug for many years. On the other hand, one of the most widely distributed mycotoxin contaminating food and animal feed is deoxynivalenol (DON. Deoxynivalenol has adverse effects on humans, animals, and crops that result in illnesses. The aim of the in vitro study was to investigated the effect of natural substance amygdalin at the selected doses (1, 10, 100, 1000, 10 000 µg/mL in combination with deoxynivalenol (1000 ng/mL on secretion of steroid hormones (progesterone and estradiol by ovarian granulosa cells (GCs from cyclic pigs. Our results showed that the releasing of progesterone and estradiol by ovarian granulosa cells was affected by amygdalin plus DON addition. The secretion of progesterone by ovarian GCs was significantly (P≤0.05 affected by administration of both compounds in all experimental groups. Similarly, estradiol releasing by GCs was significantly (P≤0.05 increased in experimental groups with amygdalin (10, 100 and 10 000 µg/mL plus DON (1000 ng/mL addition. Amygdalin treatment combined with DON caused increase of steroid hormones release by ovarian granulosa cells. Our findings suggest possible involvement of these natural substances (amygdalin and deoxynivalenol in the regulation process of steroidogenesis. In conclusion, results from this experiment contribute to knowledge about interaction between two different natural compounds and their positive or negative interferences with ovarian functions.

  4. Analytical validation of an ultraviolet-visible procedure for determining lutein concentration and application to lutein-loaded nanoparticles.

    Science.gov (United States)

    Silva, Jéssica Thaís do Prado; Silva, Anderson Clayton da; Geiss, Julia Maria Tonin; de Araújo, Pedro Henrique Hermes; Becker, Daniela; Bracht, Lívia; Leimann, Fernanda Vitória; Bona, Evandro; Guerra, Gustavo Petri; Gonçalves, Odinei Hess

    2017-09-01

    Lutein is a carotenoid presenting known anti-inflammatory and antioxidant properties. Lutein-rich diets have been associated with neurological improvement as well as reduction of the risk of vision loss due to Age-Related Macular Degeneration (AMD). Micro and nanoencapsulation have demonstrated to be effective techniques in protecting lutein against degradation and also in improving its bioavailability. However, actual lutein concentration inside the capsules and encapsulation efficiency are key parameters that must be precisely known when designing in vitro and in vivo tests. In this work an analytical procedure was validated for the determination of the actual lutein content in zein nanoparticles using ultraviolet-visible spectroscopy. Method validation followed the International Conference on Harmonisation (ICH) guidelines which evaluate linearity, detection limit, quantification limit, accuracy and precision. The validated methodology was applied to characterize lutein-loaded nanoparticles. Copyright © 2017 Elsevier Ltd. All rights reserved.

  5. Flow cytometric DNA ploidy analysis of ovarian granulosa cell tumors

    NARCIS (Netherlands)

    D. Chadha; C.J. Cornelisse; A. Schabert (A.)

    1990-01-01

    textabstractAbstract The nuclear DNA content of 50 ovarian tumors initially diagnosed as granulosa cell tumors was measured by flow cytometry using paraffin-embedded archival material. The follow-up period of the patients ranged from 4 months to 19 years. Thirty-eight tumors were diploid or near-dip

  6. Lutein transport by Caco-2 TC-7 cells occurs partly by a facilitated process involving the scavenger receptor class B type I (SR-BI).

    Science.gov (United States)

    Reboul, Emmanuelle; Abou, Lydia; Mikail, Céline; Ghiringhelli, Odette; André, Marc; Portugal, Henri; Jourdheuil-Rahmani, Dominique; Amiot, Marie-Josèphe; Lairon, Denis; Borel, Patrick

    2005-04-15

    The carotenoid lutein is thought to play a role in the human eye and to protect against age-related macular degeneration. Lutein transport in the human intestine has not been characterized. We examined lutein transport processes using Caco-2 TC-7 monolayers as a model for human intestinal epithelium. Purified lutein was mixed with phospholipids, lysophospholipids, cholesterol, mono-olein, oleic acid and taurocholate to obtain lutein-rich mixed micelles that mimicked those found under physiological conditions. The micelles were added to the apical side of Caco-2 TC-7 cell monolayers for 30 min or 3 h at 37 degrees C. Absorbed lutein, i.e. the sum of lutein recovered in the scraped cells and in the basolateral chamber, was quantified by HPLC. Transport rate was measured (i) as a function of time (from 15 to 60 min), (ii) as a function of micellar lutein concentration (from 1.5 to 15 microM), (iii) at 4 degrees C, (iv) in the basolateral to apical direction, (v) after trypsin pretreatment, (vi) in the presence of beta-carotene and/or lycopene, (vii) in the presence of increasing concentrations of antibody against SR-BI (scavenger receptor class B type 1) and (viii) in the presence of increasing concentrations of a chemical inhibitor of the selective transfer of lipids mediated by SR-BI, i.e. BLT1 (blocks lipid transport 1). The rate of transport of lutein as a function of time and as a function of concentration was saturable. It was significantly lower at 4 degrees C than at 37 degrees C (approx. 50%), in the basal to apical direction than in the opposite direction (approx. 85%), and after trypsin pretreatment (up to 45%). Co-incubation with beta-carotene, but not lycopene, decreased the lutein absorption rate (approx. 20%) significantly. Anti-SR-BI antibody and BLT1 significantly impaired the absorption rate (approx. 30% and 57% respectively). Overall, these results indicate that lutein absorption is, at least partly, protein-mediated and that some lutein is taken up

  7. Novel Lutein Loaded Lipid Nanoparticles on Porcine Corneal Distribution

    Directory of Open Access Journals (Sweden)

    Chi-Hsien Liu

    2014-01-01

    Full Text Available Topical delivery has the advantages including being user friendly and cost effective. Development of topical delivery carriers for lutein is becoming an important issue for the ocular drug delivery. Quantification of the partition coefficient of drug in the ocular tissue is the first step for the evaluation of delivery efficacy. The objectives of this study were to evaluate the effects of lipid nanoparticles and cyclodextrin (CD on the corneal lutein accumulation and to measure the partition coefficients in the porcine cornea. Lipid nanoparticles combined with 2% HPβCD could enhance lutein accumulation up to 209.2±18 (μg/g which is 4.9-fold higher than that of the nanoparticles. CD combined nanoparticles have 68% of drug loading efficiency and lower cytotoxicity in the bovine cornea cells. From the confocal images, this improvement is due to the increased partitioning of lutein to the corneal epithelium by CD in the lipid nanoparticles. The novel lipid nanoparticles could not only improve the stability and entrapment efficacy of lutein but also enhance the lutein accumulation and partition in the cornea. Additionally the corneal accumulation of lutein was further enhanced by increasing the lutein payload in the vehicles.

  8. 人卵巢颗粒细胞FoxO基因和卵泡刺激素受体基因的表达及相关性研究%EXPRESSION AND RELATIONSHIP OF FoxO GENE AND FOLLICLE STIMULATING HORMONE RECEPTOR GENE IN HUMAN OVARIAN GRANULOSA CELLS

    Institute of Scientific and Technical Information of China (English)

    梁莹; 米梅艳; 李淑贤; 周莉; 吴晓茜; 雷灵梅

    2014-01-01

    Objective To explore the expression of forkhead box(FoxO)gene and follicle-stimulating hormone receptor (FSHR)in human ovarian granulosa cells and their relationship. Methods One hundred and twenty-six unfertility women treated with in vitro fertilization and embryo transplantation (IVF-ET ) were recruited.According to the pregnancy outcome,the patients were divided into pregnancy groups 63 cases and non-pregnancy group 63 cases.The expressions of FoxO1 mRNA,FoxO3a mRNA and FSHR mRNA on human ovarian granulosa cells were detected by real time fluorescence quantitative PCR.The correlation analysis of FoxO1, FoxO3a,FSHR with serum sex hormones including luteotropic hormone(LH),estradiol(E2 ), progesterone(P)were mevaluated.Results High quality embryo number and serum E2 level in pregnancy group were significantly higher than those of non-pregnancy group (P0.05). The expression FoxO1 mRNA in pregnancy group was higher than that of non-pregnancy group (P0.05).The correlation coefficients of FoxO1 and FoxO3a with FSHR were 0.881,0.999 (P<0.01 ).Conclusion FoxO1 and FoxO3a genes participate in the regulation of oocyte quality and their functions have closely connected with FSHR.%目的:研究人卵巢颗粒细胞叉头框(forkhead box,Fox)基因和卵泡刺激素受体(follicle stimulating hormone receptor,FSHR)的表达及相关性。方法接受体外受精-胚胎移植(in vitro fertilization-embryo transfer, IVF-ET)不孕症患者126例,按照妊娠结果分为妊娠组63例,未妊娠组63例。回顾性分析取卵日成熟卵泡壁颗粒细胞 FoxO1、FoxO3a 和 FSHR mRNA 的表达,并对 FoxO1、FoxO3a、FSHR 之间及与血清促黄体生成素(luteotropic hormone,LH)、雌二醇(estradiol,E2)、孕酮(progesterone,P)进行相关性分析。结果妊娠组血清 E2水平高于未妊娠组(P<0.01);妊娠组优质胚胎数多于未妊娠组(P<0.01);2组 LH、P、获卵数差异无统计学意义(P>0

  9. Bioconversion of lutein to form aroma compounds by Pantoea dispersa.

    Science.gov (United States)

    Zhao, Yue; Zhong, Gui Fang; Yang, Xue Peng; Hu, Xian Mei; Mao, Duo Bin; Ma, Yu Ping

    2015-08-01

    To investigate the conversion of lutein, a carotenoid, to aroma compounds by Pantoea dispersa Y08, a lutein-degrading bacterium isolated from marigold flower residue. Bioconversion conditions, including substrate concentration, applied co-solvent and reaction time, were optimized. A maximum biodegradation yield of 80 % for lutein at 10 g/l was achieved. The intermediate, 3-hydroxy-β-ionone, and final β-ionone products were revealed by GC-MS. A bioconversion pathway of lutein is proposed to involve cleavage at the 9-10 double bond position, followed by de-hydroxylation at the 3-hydroxy position. This is the first report of the ability of a bacterium, P. dispersa, to sequentially convert lutein to 3-hydroxy-β-ionone and then β-ionone.

  10. Transforming growth factor alpha (TGFα regulates granulosa cell tumor (GCT cell proliferation and migration through activation of multiple pathways.

    Directory of Open Access Journals (Sweden)

    Cheng Wang

    Full Text Available Granulosa cell tumors (GCTs are the most common ovarian estrogen producing tumors, leading to symptoms of excessive estrogen such as endometrial hyperplasia and endometrial adenocarcinoma. These tumors have malignant potential and often recur. The etiology of GCT is unknown. TGFα is a potent mitogen for many different cells. However, its function in GCT initiation, progression and metastasis has not been determined. The present study aims to determine whether TGFα plays a role in the growth of GCT cells. KGN cells, which are derived from an invasive GCT and have many features of normal granulosa cells, were used as the cellular model. Immunohistochemistry, Western blot and RT-PCR results showed that the ErbB family of receptors is expressed in human GCT tissues and GCT cell lines. RT-PCR results also indicated that TGFα and EGF are expressed in the human granulosa cells and the GCT cell lines, suggesting that TGFα might regulate GCT cell function in an autocrine/paracrine manner. TGFα stimulated KGN cell DNA synthesis, cell proliferation, cell viability, cell cycle progression, and cell migration. TGFα rapidly activated EGFR/PI3K/Akt and mTOR pathways, as indicated by rapid phosphorylation of Akt, TSC2, Rictor, mTOR, P70S6K and S6 proteins following TGFα treatment. TGFα also rapidly activated the EGFR/MEK/ERK pathway, and P38 MAPK pathways, as indicated by the rapid phosphorylation of EGFR, MEK, ERK1/2, P38, and CREB after TGFα treatment. Whereas TGFα triggered a transient activation of Akt, it induced a sustained activation of ERK1/2 in KGN cells. Long-term treatment of KGN cells with TGFα resulted in a significant increase in cyclin D2 and a decrease in p27/Kip1, two critical regulators of granulosa cell proliferation and granulosa cell tumorigenesis. In conclusion, TGFα, via multiple signaling pathways, regulates KGN cell proliferation and migration and may play an important role in the growth and metastasis of GCTs.

  11. Evaluation of the simultaneous production of lutein and lipids using a vertical alveolar panel bioreactor for three Chlorella species 

    OpenAIRE

    2014-01-01

    The concept of a biorefinery improves the economic efficiency of a biofuel production process from microalgae by recovering high value added compounds. Lutein is a carotenoid currently extracted from petals of Tagetes erecta with an established market in poultry and in human nutritional supplements. For the very first time, an extended study on the lipid and lutein production over three Chlorella species as well as cell disruption methods was performed. Chlorella vulgaris, Chlorella zofingien...

  12. Changes in Macular Pigment Optical Density and Serum Lutein Concentration in Japanese Subjects Taking Two Different Lutein Supplements.

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    Akira Obana

    Full Text Available To investigate macular pigment optical density (MPOD and serum concentration changes of lutein in Japanese subjects participating in a clinical trial in which two formulations of lutein and zeaxanthin supplements with different physiochemical properties are used.Thirty-six healthy volunteers were recruited into this prospective, randomized, parallel-group, double-masked comparative study at a single institute. Two products were used, FloraGLO® (Kemin Japan and XanMax® (Katra Phytochem. The lutein particle size and zeaxanthin concentrations differed between the formulations. The subjects consumed one of the two supplements for a duration of up to 6 months. MPOD levels were measured by resonance Raman spectrometry at baseline and once a month until the end of the study. Serum lutein concentration was measured at baseline, month 3, and month 6. The subjects were also tested for contrast sensitivity, glare sensitivity, visual acuity, and in addition had a focal electroretinogram measured.The mean serum lutein concentrations increased significantly after the first three months, but the mean MPOD levels in either supplement group did not show any statistically significant increase. A detailed analysis, however, revealed three response patterns in both groups for the increase of MPOD levels and serum lutein concentration, i.e. "retinal responders", who had an increase of both MPOD levels and serum lutein concentrations (n = 13, "retinal non-responders", who had only increased serum concentrations and no change in MPOD levels (n = 20, and "retinal and serum non-responders", who had neither MPOD level nor plasma concentration increases (n = 3. The subjects with low MPOD levels at baseline appeared to show increased MPOD levels at the 6 month time point upon lutein supplementation (r = -0.4090, p = 0.0133. Glare sensitivity improved in retinal responders in both supplement groups, while there were no remarkable changes in contrast sensitivity

  13. FSH regulates acetycholine production by ovarian granulosa cells

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    Amsterdam Abraham

    2006-07-01

    Full Text Available Abstract Background It has been previously shown that cultured granulosa cells (GCs derived from human ovarian preovulatory follicles contain choline acetyltransferase (ChAT, the enzyme responsible for acetylcholine (ACh synthesis. They also produce ACh and express functional muscarinic ACh receptors. ACh can act on GCs to increase proliferation, disrupt gap junctional communication, alter intracellular calcium levels, as well as expression of transcription factors, suggesting an unrecognized role of ACh in GC function. To gain further insights into the possible role of ACh in the ovary, we examined ChAT expression in the gland before and after birth, as well as in adults, and studied the regulation of ACh production by FSH. Methods ChAT immunohistochemistry was performed using ovarian samples of different species and ages (embryonic, postnatal and adult rats and mice, including embryonic ovaries from mice null for ChAT, neonatal and adult rhesus monkeys and adult humans. ACh was measured by HPLC and/or a fluorescence based method in rat ovaries and in a FSH receptor-expressing cell line (rat GFSHR-17 cultured with or without FSH. Results In adult rat, as well as in all other species, ovarian ChAT immunoreactivity is associated with GCs of antral follicles, but not with other structures, indicating that GCs are the only ovarian source of ACh. Indeed ACh was clearly detected in adult rat ovaries by two methods. ChAT immunoreactivity is absent from embryonic and/or neonatal ovaries (mouse/rat and monkey and ovarian development in embryonic mice null for ChAT appears normal, suggesting that ACh is not involved in ovarian or follicular formation. Since ChAT immunoreactivity is present in GCs of large follicles and since the degree of the ChAT immunoreactivity increases as antral follicles grow, we tested whether ACh production is stimulated by FSH. Rat GFSHR-17 cells that stably express the FSH receptor, respond to FSH with an increase in ACh

  14. Effects of Melatonin on the Proliferation and Apoptosis of Sheep Granulosa Cells under Thermal Stress

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    Yao Fu

    2014-11-01

    Full Text Available The cross-talk between oocyte and somatic cells plays a crucial role in the regulation of follicular development and oocyte maturation. As a result, granulosa cell apoptosis causes follicular atresia. In this study, sheep granulosa cells were cultured under thermal stress to induce apoptosis, and melatonin (MT was examined to evaluate its potential effects on heat-induced granulosa cell injury. The results demonstrated that the Colony Forming Efficiency (CFE of granulosa cells was significantly decreased (heat 19.70% ± 1.29% vs. control 26.96% ± 1.81%, p < 0.05 and the apoptosis rate was significantly increased (heat 56.16% ± 13.95% vs. control 22.80% ± 12.16%, p < 0.05 in granulosa cells with thermal stress compared with the control group. Melatonin (10−7 M remarkably reduced the negative effects caused by thermal stress in the granulosa cells. This reduction was indicated by the improved CFE and decreased apoptotic rate of these cells. The beneficial effects of melatonin on thermal stressed granulosa cells were not inhibited by its membrane receptor antagonist luzindole. A mechanistic exploration indicated that melatonin (10−7 M down-regulated p53 and up-regulated Bcl-2 and LHR gene expression of granulosa cells under thermal stress. This study provides evidence for the molecular mechanisms of the protective effects of melatonin on granulosa cells during thermal stress.

  15. Effect of dietary supplementation of different lutein sources on the productive performance of commercial layers categorized in three body weights

    Directory of Open Access Journals (Sweden)

    A.A. Sajjad

    2013-06-01

    Full Text Available Lutein enrichment can be used for reducing chances of age related macular degeneration (AMD in human being. The present study was conducted to evaluate the production performance of commercial layers categorized in three body weight categories supplemented with different lutein sources. For this purpose a total number of four thirty two 31 week old Hy-line (W-36 commercial layers were categorized in three body sizes (Heavy >1400g, Medium 1300-1399 g, Light <1300 and supplemented with different sources of Lutein (Control, Free, Esterified and Free + Esterified replicated 6 times containing 6 birds each. The data were collected for production performance (egg production %, egg weight, egg mass, FCR per dozen, FCR per kg egg mass. The data were analyzed using analysis of variance (ANOVA technique through Completely Randomized Design (CRD. Results of the present study showed non-significant (P>0.05 differences for production performance among different lutein sources and body weight categories except egg weight which was found to be highest in heavy birds. Thus Lutein can be supplemented in layer diet without any harm to its production performance.

  16. Leptin siRNA promotes ovarian granulosa cell apoptosis and affects steroidogenesis by increasing NPY2 receptor expression.

    Science.gov (United States)

    Ding, Xiaomeng; Kou, Xinxin; Zhang, Ye; Zhang, Xiaoli; Cheng, Guomei; Jia, Tianming

    2017-10-30

    Leptin has been found to be involved in the ovarian granulosa cell apoptosis and steroidogenesis. Loss of neuropeptide Y (NPY) can correct the obesity syndrome of mutant mice lacking of leptin (ob/ob). However, the association of NPY and leptin in ovarian granulosa cells and ovarian steroidogenesis has not been investigated. Here, C57BL/6J ob/ob mice and C57BL/6J (control) mice were intraperitoneally injected with PBS, leptin (0.4μg/g bodyweight) or BIIE0246 (NPY2 receptor [NPY2R] antagonist, 30μg/kg bodyweight) every day for 15days. We found that NPY2R mRNA expression in mouse ovary was suppressed by leptin treatment, but increased by leptin deficiency. Leptin or BIIE0246 treatment significantly increased E2, but notably decreased progesterone in both mice. A lower level of E2 and a higher level of progesterone was observed in ob/ob mice than in control mice. Further, we then knocked down leptin expression in human ovarian granulosa cells by siRNA transfection and treated the cells with DMSO or BIIE0246. In vitro experiments confirmed the findings in mice. siLeptin treatment decreased the secretion of E2, anti-Mullerian hormone (AMH), insulin-like growth factor (IGF)-1 and transforming growth factor (TGF)-β, and the cell proliferation, but increased the secretion of progesterone and cell apoptosis. Western blotting analysis of PCNA, Bcl-2 and Bax confirmed the results of cell proliferation and apoptosis. Activation of JAK2 and STAT3 was also suppressed by knocking down leptin. All the effects of siLeptin on ovarian granulosa cells were partially reversed by BIIE0246. In conclusion, knockdown of leptin significantly affected ovarian steroidogenesis and ovarian function through NPY. siLeptin transfection impaired the activation of JAK2/STAT3 and contributed to ovarian granulosa cell apoptosis partially through up-regulating NPY2R expression. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Vasoactive intestinal peptide-induced expression of cytochrome P450 cholesterol side-chain cleavage and 17 alpha-hydroxylase enzyme activity in hen granulosa cells.

    Science.gov (United States)

    Johnson, A L; Li, Z; Gibney, J A; Malamed, S

    1994-08-01

    Experiments were conducted to determine whether vasoactive intestinal peptide (VIP) can regulate expression of cytochrome P450 side-chain cleavage (P450scc) and P450 17 alpha-hydroxylase (P450 17 alpha-OH) mRNA levels and enzyme activity in granulosa cells from nonhierarchal (6-8-mm) follicles. Initial studies demonstrated that immunoreactive VIP is localized within the theca (but not granulosa) layer of both resting (< 0.5-mm follicles) and 6-8-mm follicles, thus providing a potential paracrine mechanism of action for VIP. While short-term (3 h) incubation of granulosa cells with VIP (0.001-1.0 microM) failed to stimulate progesterone production from 6-8-mm follicle granulosa cells, a 4-h culture period in the presence of VIP resulted in increased cyclic AMP (cAMP) accumulation, and a 24-h culture period resulted in progesterone synthesis and increased P450scc mRNA levels; control levels of each endpoint measurement were not altered within the period observed. By contrast, culture with the growth factor transforming growth factor alpha (TGF alpha) in the presence of VIP (1 microM) prevented increases in P450scc mRNA levels and progesterone production. Similar effects of VIP and TGF alpha in the presence of VIP were demonstrated for P450 17 alpha-OH mRNA levels and enzyme activity. Finally, there was an additive effect of VIP (0.1 microM) plus recombinant human (rh) FSH (100 mIU) on the initiation of progesterone production in cultured 6-8-mm follicle granulosa cells compared to the addition of VIP or rhFSH alone.(ABSTRACT TRUNCATED AT 250 WORDS)

  18. Induction of follicular luteinization by equine chorionic gonadotropin in cyclic guinea pigs

    Institute of Scientific and Technical Information of China (English)

    Jun-rong LI; Wei WANG; Fang-xiong SHI

    2015-01-01

    题目:正常发情周期豚鼠经孕马血清促性腺激素诱导的卵泡黄体化研究  目的:研究孕马血清促性腺激素(eCG)对发情周期豚鼠卵巢卵泡的作用。  创新点:首次发现eCG对于发情周期豚鼠发挥了类似促黄体素的作用,而非促卵泡素的作用。  方法:将成年雌性豚鼠(400~700 g,连续2次以上观察到稳定的16天发情周期)分为对照组(腹腔注射生理盐水)和实验组(腹腔注射eCG)。实验组根据注射强度分为20 IU组和50 IU组,并分别于注射后4和8天采集豚鼠卵巢。用苏木精-伊红染色法(H&E)和免疫组化法观察豚鼠卵巢变化情况。测定注射后4天卵巢卵泡大小和数量,测定注射后8天卵巢和子宫重量、黄体数量、黄体化细胞数量和闭锁黄体细胞比例。  结论:本实验中,豚鼠经eCG注射后卵巢变化结果显示:发情期豚鼠卵巢经 eCG 注射发生明显的形态改变(图1),50 IU组豚鼠卵巢在注射后8天出现了黄体化未破裂卵泡(LUF)现象(图3)。免疫组化结果显示增值细胞核抗原(PCNA)和类激素调节蛋白(StAR)都免疫定位于黄体化卵泡(图8)。综上所述,eCG 对于发情期豚鼠发挥了类似促黄体素的作用。%The effects of equine chorionic gonadotropin (eCG) on follicular development and ovulation in cyclic guinea pigs were investigated by histological and immunohistochemical analyses. Three groups of guinea pigs (n=12) were administrated subcutaneously with saline, 20 or 50 IU of eCG, respectively, on cyclic Day 12 (Day 1=vaginal openings). Ovaries were collected at 4 and 8 d after administration (6 animals per group each time). The eCG ad-ministration induced significant and distinct morphological changes in the ovaries, as it promoted the luteinization of granulosa cel s, but not fol icular development. In addition, proliferating cell nuclear antigen (PCNA) and

  19. Juvenile granulosa cell tumour: a rare clinical entity

    Directory of Open Access Journals (Sweden)

    Kaliki Hymavathi Reddy

    2014-08-01

    Full Text Available Ovarian cancer is the third most common neoplasm of the female genital tract. Based on the cell type of origin, primary ovarian malignancies are classified into surface epithelium, germ cell, and sex cord tumors. Sex cord tumors account for 1% to 2% of ovarian malignancies. They may contain granulosa cells, theca cells, sertoli cells, or fibroblasts of gonadal stromal origin. Granulosa Cell Tumours (GCTs account for approximately 2-5% of all ovarian tumors and can be divided into adult (95% and juvenile (5% types based on histologic findings. GCTs secrete estrogen thus resulting in menstrual irregularities in the affected individual. More serious estrogen effects can occur in various end organs such as uterus resulting in endometrial hyperplasia, endometrial adenocarcinomas and increased risk of breast cancers. Androgen production is also reported but rare and produces virilization in the affected women. Juvenile Granulosa Cell Tumours (JGCTs are clinically and histopathologically distinct from the GCTs. They are rarely encountered but mostly in youngsters. Surgery is the primary modality of treatment with chemotherapy being reserved for advanced or recurrent disease states. We herewith report an interesting case of JGCT in a young teenage girl. [Int J Reprod Contracept Obstet Gynecol 2014; 3(4.000: 1150-1154

  20. Comparison of lutein bioavailability from vegetables and supplement.

    Science.gov (United States)

    Riso, Patrizia; Brusamolino, Antonella; Ciappellano, Salvatore; Porrini, Marisa

    2003-05-01

    Lutein is a carotenoid present in dark green leafy vegetables and it may be involved in the prevention of several diseases related to oxidative stress. The aim of this study was to evaluate bioavailability of lutein from different food sources (150 g spinach and 200 g broccoli) and a supplement in oil (300 mg VEGEX), all providing about 9 mg lutein. Eight healthy females were instructed to eat a low-carotenoid diet for the period of experimentation. On three different occasions, three weeks apart, volunteers ate the lutein sources together with 10 g olive oil and 40 g bread. Blood samples were collected just before eating, every two hours for 12 hours, and at 24, 32, 56, 80 and 104 hours. Lutein concentration increased significantly after six to eight hours and peaked after 10-12 hours, with the highest concentration reached after VEGEX intake. Lutein concentration remained significantly elevated for up to 80 hours (VEGEX and spinach). On the whole, our results suggest that the intake of one single dose of lutein from different sources is able to bring about a significant plasma response in the short term.

  1. Administration of follicle-stimulating hormone induces autophagy via upregulation of HIF-1[alpha] in mouse granulosa cells

    National Research Council Canada - National Science Library

    Jilong Zhou; Wang Yao; Chengyu Li; Wangjun Wu; Qifa Li; Honglin Liu

    2017-01-01

    ...) on mouse granulosa cells (MGCs). Results indicated that autophagy was induced by FSH, which is known to be the dominant hormone regulating follicular development and granulosa cell (GC) proliferation...

  2. Mural granulosa cell gene expression associated with oocyte developmental competence

    Directory of Open Access Journals (Sweden)

    Jiang Jin-Yi

    2010-03-01

    Full Text Available Abstract Background Ovarian follicle development is a complex process. Paracrine interactions between somatic and germ cells are critical for normal follicular development and oocyte maturation. Studies have suggested that the health and function of the granulosa and cumulus cells may be reflective of the health status of the enclosed oocyte. The objective of the present study is to assess, using an in vivo immature rat model, gene expression profile in granulosa cells, which may be linked to the developmental competence of the oocyte. We hypothesized that expression of specific genes in granulosa cells may be correlated with the developmental competence of the oocyte. Methods Immature rats were injected with eCG and 24 h thereafter with anti-eCG antibody to induce follicular atresia or with pre-immune serum to stimulate follicle development. A high percentage (30-50%, normal developmental competence, NDC of oocytes from eCG/pre-immune serum group developed to term after embryo transfer compared to those from eCG/anti-eCG (0%, poor developmental competence, PDC. Gene expression profiles of mural granulosa cells from the above oocyte-collected follicles were assessed by Affymetrix rat whole genome array. Results The result showed that twelve genes were up-regulated, while one gene was down-regulated more than 1.5 folds in the NDC group compared with those in the PDC group. Gene ontology classification showed that the up-regulated genes included lysyl oxidase (Lox and nerve growth factor receptor associated protein 1 (Ngfrap1, which are important in the regulation of protein-lysine 6-oxidase activity, and in apoptosis induction, respectively. The down-regulated genes included glycoprotein-4-beta galactosyltransferase 2 (Ggbt2, which is involved in the regulation of extracellular matrix organization and biogenesis. Conclusions The data in the present study demonstrate a close association between specific gene expression in mural granulosa cells and

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  11. Action of luteinizing hormone-releasing hormone in rat ovarian cells: Hormone production and signal transduction

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Jian.

    1989-01-01

    The present study was conducted to investigate the hypothesis that the breakdown of membrane phosphoinositides may participate in the actions of luteinizing hormone-releasing hormone (LHRH) on hormone production in rat granulosa cells. In cells prelabeled with ({sup 3}H)inositol or ({sup 3}H)arachidonic acid (AA), treatment with LHRH increased the formation of radiolabeled inositol 1,4,5-trisphosphate (IP{sub 3}) and diacylglycerol (DG), and the release of radiolabeled AA. Since IP{sub 3} induces intracellular Ca{sup 2+} mobilization, changes in the cytosolic free calcium ion concentrations ((Ca{sup 2+})i) induced by LHRH were studied in individual cells using fura-2 microspectrofluorimetry. Alterations in (Ca{sup 2+})i induced by LHRH were rapid and transient, and could be completely blocked by a LHRH antagonist. Sustained perifusion of LHRH resulted in a desensitization of the (Ca{sup 2+})i response to LHRH. LHRH treatment accelerated (Ca{sup 2+})i depletion in the cells perifused with Ca{sup 2+} free medium, indicating the involvement of intracellular Ca{sup 2+} pool(s) in (Ca{sup 2+})i changes. The actions of LHRH on the regulation of progesterone (P{sub 4}) and prostaglandin E{sub 2} (PGE{sub 2}) production were also examined. LHRH increased basal P{sub 4} production and attenuated FSH induced P{sub 4} production. Both basal and FSH stimulated PGE{sub 2} formation were increased by LHRH. Since LHRH also increased the formation of DG that stimulates the activity of protein kinase C, an activator of protein kinase C (12-0-tetradecanolyphorbol-13-acetate: TPA) was used with the Ca{sup 2+} ionophore A23187 and melittin (an activator of phospholipase A{sub 2}) to examine the roles of protein kinase C, Ca{sup 2+} and free AA, respectively, in LHRH action.

  12. Chemokine Ligand 20: A Signal for Leukocyte Recruitment During Human Ovulation?

    Science.gov (United States)

    Al-Alem, Linah; Puttabyatappa, Muraly; Rosewell, Kathy; Brännström, Mats; Akin, James; Boldt, Jeffrey; Muse, Ken; Curry, Thomas E

    2015-09-01

    Ovulation is one of the cornerstones of female fertility. Disruption of the ovulatory process results in infertility, which affects approximately 10% of couples. Using a unique model in which the dominant follicle is collected across the periovulatory period in women, we have identified a leukocyte chemoattractant, chemokine ligand 20 (CCL20), in the human ovary. CCL20 mRNA is massively induced after an in vivo human chorionic gonadotropin (hCG) stimulus in granulosa (>10 000-fold) and theca (>4000-fold) cells collected during the early ovulatory (12-18 h) and late ovulatory (18-34 h) periods after hCG administration. Because the LH surge sets in motion an inflammatory reaction characterized by an influx of leukocytes and CCL20 is known to recruit leukocytes in other systems, the composition of ovarian leukocytes (CD45+) containing the CCL20 receptor CCR6 was determined immediately prior to ovulation. CD45+/CCR6+ cells were primarily natural killer cells (41%) along with B cells (12%), T cells (11%), neutrophils (10%), and monocytes (9%). Importantly, exogenous CCL20 stimulated ovarian leukocyte migration 59% within 90 minutes. Due to the difficulties in obtaining human follicles, an in vitro model was developed using granulosa-lutein cells to explore CCL20 regulation. CCL20 expression increased 40-fold within 6 hours after hCG, was regulated partially by the epithelial growth factor pathway, and was positively correlated with progesterone production. These results demonstrate that hCG dramatically increases CCL20 expression in the human ovary, that ovarian leukocytes contain the CCL20 receptor, and that CCL20 stimulates leukocyte migration. Our findings raise the prospect that CCL20 may aid in the final ovulatory events and contribute to fertility in women.

  13. Lutein Inhibits the Migration of Retinal Pigment Epithelial Cells via Cytosolic and Mitochondrial Akt Pathways (Lutein Inhibits RPE Cells Migration

    Directory of Open Access Journals (Sweden)

    Ching-Chieh Su

    2014-08-01

    Full Text Available During the course of proliferative vitreoretinopathy (PVR, the retinal pigment epithelium (RPE cells will de-differentiate, proliferate, and migrate onto the surfaces of the sensory retina. Several studies have shown that platelet-derived growth factor (PDGF can induce migration of RPE cells via an Akt-related pathway. In this study, the effect of lutein on PDGF-BB-induced RPE cells migration was examined using transwell migration assays and Western blot analyses. We found that both phosphorylation of Akt and mitochondrial translocation of Akt in RPE cells induced by PDGF-BB stimulation were suppressed by lutein. Furthermore, the increased migration observed in RPE cells with overexpressed mitochondrial Akt could also be suppressed by lutein. Our results demonstrate that lutein can inhibit PDGF-BB induced RPE cells migration through the inhibition of both cytoplasmic and mitochondrial Akt activation.

  14. Regulation of granulosa cell cocaine and amphetamine regulated transcript (CART) binding and effect of CART signaling inhibitor on granulosa cell estradiol production during dominant follicle selection in cattle.

    Science.gov (United States)

    Folger, Joseph K; Jimenez-Krassel, Fermin; Ireland, James J; Lv, Lihua; Smith, George W

    2013-12-01

    We previously established a potential role for cocaine and amphetamine regulated transcript (CARTPT) in dominant follicle selection in cattle. CARTPT expression is elevated in subordinate versus dominant follicles, and treatment with the mature form of the CARTPT peptide (CART) decreases follicle-stimulating hormone (FSH)-stimulated granulosa cell estradiol production in vitro and follicular fluid estradiol and granulosa cell CYP19A1 mRNA in vivo. However, mechanisms that regulate granulosa cell CART responsiveness are not understood. In this study, we investigated hormonal regulation of granulosa cell CART-binding sites in vitro and temporal regulation of granulosa cell CART-binding sites in bovine follicles collected at specific stages of a follicular wave. We also determined the effect of inhibition of CART receptor signaling in vivo on estradiol production in future subordinate follicles. Granulosa cell CART binding in vitro was increased by FSH, and this induction was blocked by estrogen receptor antagonist treatment. In follicles collected in vivo at specific stages of a follicular wave, granulosa cell CART binding in the F2 (second largest), future subordinate follicle increased during dominant follicle selection. Injection into the F2 follicle (at onset of diameter deviation) of an inhibitor of the o/i subclass of G proteins (previously shown to block CART actions in vitro) resulted in increased follicular fluid estradiol concentrations in vivo. Collectively, results demonstrate hormonal regulation of granulosa cell CART binding in vitro and temporal regulation of CART binding in subordinate follicles during dominant follicle selection. Results also suggest that CART signaling may help suppress estradiol-producing capacity of the F2 (subordinate) follicle during this time period.

  15. Stimulatory Effect of Insulin on 5α-Reductase Type 1 (SRD5A1) Expression through an Akt-Dependent Pathway in Ovarian Granulosa Cells

    Science.gov (United States)

    Kayampilly, Pradeep P.; Wanamaker, Brett L.; Stewart, James A.; Wagner, Carrie L.; Menon, K. M. J.

    2010-01-01

    Elevated levels of 5α-reduced androgens have been shown to be associated with hyperandrogenism and hyperinsulinemia, the leading causes of ovulatory dysfunction in women. 5α-Dihydrotestosterone reduces ovarian granulosa cell proliferation by inhibiting FSH-mediated mitogenic signaling pathways. The present study examined the effect of insulin on 5α-reductase, the enzyme that catalyses the conversion of androgens to their 5α-derivatives. Granulosa cells isolated from immature rat ovaries were cultured in serum-free, phenol red-free DMEM-F12 media and treated with different doses of insulin (0, 0.1, 1.0, and 10.0 μg/ml) for different time intervals up to 12 h. The expression of 5α-reductase type 1 mRNA, the predominant isoform found in granulosa cells, showed a significant (P < 0.05) increase in response to the insulin treatment up to 12 h compared with control. The catalytic activity of 5α-reductase enzyme was also stimulated in a dose-depended manner (P < 0.05). Inhibiting the Akt-dependent signaling pathway abolished the insulin-mediated increase in 5α-reductase mRNA expression, whereas inhibition of the ERK-dependent pathway had no effect. The dose-dependent increase in 5α-reductase mRNA expression as well as catalytic activity seen in response to insulin treatment was also demonstrated in the human granulosa cell line (KGN). In addition to increased mRNA expression, a dose-dependent increase in 5α-reductase protein expression in response to insulin was also seen in KGN cells, which corroborated well with that of mRNA expression. These results suggest that elevated levels of 5α-reduced androgens seen in hyperinsulinemic conditions might be explained on the basis of a stimulatory effect of insulin on 5α-reductase in granulosa cells. The elevated levels of these metabolites, in turn, might adversely affect growth and proliferation of granulosa cells, thereby impairing follicle growth and ovulation. PMID:20810561

  16. Dynamics of the membrana granulosa during expansion of the ovarian follicular antrum.

    Science.gov (United States)

    Rodgers, R J; Irving-Rodgers, H F; van Wezel, I L; Krupa, M; Lavranos, T C

    2001-01-22

    As an endocrine organ, the ovary has some unique characteristics. The formation, the maturation and the regression of the hormone producing cells really determine the timing, the amount and the type of hormone secreted. Here, we focus on the granulosa cells of ovarian follicles which express 17beta-hydroxysteroid dehydrogenase type 1 and cytochrome P450 aromatase. Follicles only produce estradiol late in follicular development before either ovulation or atresia ensues. We discuss the evidence that the membrana granulosa has many characteristics in common with other epithelia, including that it arises from stem cells. The corollary of this is that individual cells within the membrana granulosa are of different ages or stages of specialization. This is evident as regional differences across the membrana granulosa in terms of cell ages, shapes, gene expression, and even behaviour on cell death. We discuss theoretical considerations of the effects of antrum formation on the behavior of the membrana granulosa, and show evidence for differences between follicles in cell shapes, basal lamina phenotypes and location of younger cells, which we speculate is due to different rates of antrum expansion. Clearly, the membrana granulosa is dynamic, and this could explain much about the differences in the behaviors of cells from within the membrana granulosa, and between ovarian follicles.

  17. Laparoscopic ovariectomy in two horses with granulosa cell tumors.

    Science.gov (United States)

    Ragle, C A; Southwood, L L; Hopper, S A; Buote, P L

    1996-09-15

    Two mares were admitted for ovariectomy of unilateral granulosa cell tumors. Both mares were ovariectomized (1 unilateral and 1 bilateral) by use of a ventral abdominal laparoscopic technique. This approach required tilting the operative table 30 degrees to elevate the pelvis and to allow observation of the ovaries. Using a single laparoscopic portal and 3 to 4 instrument portals, a triangulation technique was used. The ovarian pedicles were isolated and secured via loop ligation. The ovaries then were divided from the ligated pedicle and placed within specimen bags for extraction. The specimen bags then were removed through a ventral midline celiotomy. Using this technique, it was determined that granulosa cell tumors or ovaries of up to 20 cm in diameter can be removed. Laparoscopic ovariectomy provided a means to provide tension-free dissection and ligation of the ovarian pedicle. In comparison to conventional techniques, this may improve suture security and reduce complications related to excessive pedicle tension. Improved observation during surgery, less pedicle tension, and minimal invasiveness made laparoscopic ovariectomy of these 2 mares advantageous.

  18. Lutein derived fragments exhibit higher antioxidant and anti-inflammatory properties than lutein in lipopolysaccharide induced inflammation in rats.

    Science.gov (United States)

    Nidhi, Bhatiwada; Sharavana, Gurunathan; Ramaprasad, Talahalli R; Vallikannan, Baskaran

    2015-02-01

    In the present study, we appraise the anti-inflammatory efficacy of lutein oxidative degradation derivatives mediated through UV-irradiation over lutein in counteracting the inflammation induced by lipopolysaccharide (LPS) in rats (n = 5 per group). UV-irradiated lutein fragments were identified as anhydrolutein (B, C40H54O), 2,6,6-trimethylcyclohexa-1,4-dienylium (M1, C9H13), (2E,4E,6E,8E)-9-(4-hydroxy-2,6,6-trimethylcyclohex-1-1en-1-yl)-3,7-dimethylnona-2,4,6,8-tetraen-1-ylium (M2, C20H29O), 4-[(1E,3E,5E,7E)-3,7,-dimethyldeca-1,3,5,7-tetraen-1-yl]-3,5,5-methylcyclohex-3-en-1-ol (M3, C21H30O) and zeaxanthin (M4, C40H56O) and its isomers as 13'-Z zeaxanthin, 13'-Z lutein, all-trans zeaxanthin, and 9-Z lutein. Induction of inflammation by LPS significantly increased the production of nitrites (3.3 fold in the serum and 2.6 fold in the liver), prostaglandin E2 (26 fold in the serum), and pro-inflammatory cytokines like tumor necrosis factor-α (6.6 fold in the serum), and interleukin-6 (4.8 fold in the serum). Oxidative derivatives of lutein, especially M1, M2 and M3, ameliorated acute inflammation in rats by inhibiting the production of nitrites, malondialdehyde (MDA), PGE2, TNF-α, and IL-6 cytokines more efficiently than lutein in rats. The anti-inflammatory mechanism of derivatives might be related to the decrease of inflammatory cytokines and the increase of antioxidant enzymes (superoxide dismutase, catalase, glutathione peroxidase, glutathione S transferase, glutathione reductase), which would result in the reduction of iNOS, COX-2 and MDA and subsequently inflammatory responses.

  19. Lutein recovery from Chlorella sp. ESP-6 with coagulants.

    Science.gov (United States)

    Utomo, Rhesa Pramudita; Chang, Yin-Ru; Lee, Duu-Jong; Chang, Jo-Shu

    2013-07-01

    Production of algal lutein included cell cultivation, biomass harvesting, cell wall disruption, and subsequent purification if needed. This work cultivated Chlorella sp. ESP-6 cells in photobioreactor to a biomass content of 1.1 gl(-1) and then the freezing-grinding, ultrasonic treatment (20 and 42kHz) and microwave treatment were used to disrupt the cell walls for recover intracellular lutein. The grinding recovered more lutein than ultrasound or microwave pretreatment. Single coagulation using >30 mgl(-1) chitosan or dual-conditioning using 10 mg l(-1) polyaluminum chloride and 10 mgl(-1) chitosan effectively enhance sedimentation and membrane filtration efficiency of algal suspensions. However, the presence of coagulants lowers the lutein yield from algal biomass in the subsequent 20 kHz ultrasound treatment and purification process. Simulation results revealed affine adsorption of lutein onto chitosan molecules via hydroxyl-amine interaction. The possible drawback by pre-treatment stage should be considered together with the subsequent recovery stage in whole process assessment.

  20. Ovarian juvenile granulosa cell tumor associated with Maffucci's syndrome:case report

    Institute of Scientific and Technical Information of China (English)

    袁键群; 林小娜; 许敬尧; 祝佳; 郑伟良

    2004-01-01

    @@ Granulosa cell tumors (GCTs) are the most common ovarian sex-cord stromal tumors. Two histopathologically well defined patterns of GCTs are known: adult granulosa cell tumor (AGCT) and juvenile granulosa cell tumor (JGCT). JGCTs are rare and those associated with enchondromatosis are much rarer. A review of the literature revealed four previous cases of JGCT were associated with Maffucci ' s syndrome (MS),1-4 and nine with Ollier ' s disease (OD). This report describes ovarian JGCT with MS in a 21-year-old woman, and reviews the clinicopathological causes of both disorders.

  1. Relationship between lutein and mycotoxin content in durum wheat.

    Science.gov (United States)

    Delgado, Rosa M; Sulyok, Michael; Jirsa, Ondřej; Spitzer, Tomáš; Krska, Rudolf; Polišenská, Ivana

    2014-01-01

    Levels of lutein and a number of mycotoxins were determined in seven varieties of durum wheat (Triticum durum) and two varieties of common wheat (Triticum aestivum) in order to explore possible relationships amongst these components. Durum wheat cultivars always showed both higher lutein and mycotoxin contents than common wheat cultivars. The mycotoxins detected in both common and durum wheat cultivars were produced by the genera Fusarium, Claviceps, Alternaria and Aspergillus. Fusarium was the major producer of mycotoxins (26 mycotoxins) followed by Claviceps (14 mycotoxins), which was present only in some cultivars such as Chevalier (common wheat), Lupidur and Selyemdur (both durum wheat), Alternaria (six mycotoxins) and Aspergillus (three mycotoxins). Positive correlations between the levels of lutein and mycotoxins in durum wheat cultivars were found for the following mycotoxins: deoxynivalenol (DON), its derivative DON-3-glucoside, moniliformin, culmorin and its derivatives (5-hydroxyculmorin and 15-hydroxyculmorin).

  2. Ovotoxic effects of galactose involve attenuation of follicle-stimulating hormone bioactivity and up-regulation of granulosa cell p53 expression.

    Directory of Open Access Journals (Sweden)

    Sayani Banerjee

    Full Text Available Clinical evidence suggests an association between galactosaemia and premature ovarian insufficiency (POI; however, the mechanism still remains unresolved. Experimental galactose toxicity in rats produces an array of ovarian dysfunction including ovarian development with deficient follicular reserve and follicular resistance to gonadotrophins that characterize the basic tenets of human POI. The present investigation explores if galactose toxicity in rats attenuates the bioactivity of gonadotrophins or interferes with their receptor competency, and accelerates the rate of follicular atresia. Pregnant rats were fed isocaloric food-pellets supplemented with or without 35% D-galactose from day-3 of gestation and continuing through weaning of the litters. The 35-day old female litters were autopsied. Serum galactose-binding capacity, galactosyltransferase (GalTase activity, and bioactivity of FSH and LH together with their receptor competency were assessed. Ovarian follicular atresia was evaluated in situ by TUNEL. The in vitro effects of galactose were studied in isolated whole follicles in respect of generation of reactive oxygen species (ROS and expression of caspase 3, and in isolated granulosa cells in respect of mitochondrial membrane potential, expression of p53, and apoptosis. The rats prenatally exposed to galactose exhibited significantly decreased serum GalTase activity and greater degree of galactose-incorporation capacity of sera proteins. LH biopotency and LH-FSH receptor competency were comparable between the control and study population, but the latter group showed significantly attenuated FSH bioactivity and increased rate of follicular atresia. In culture, galactose increased follicular generation of ROS and expression of caspase 3. In isolated granulosa cells, galactose disrupted mitochondrial membrane potential, stimulated p53 expression, and induced apoptosis in vitro; however co-treatment with either FSH or estradiol

  3. Gene expression patterns in granulosa cells and oocytes at various stages of follicle development as well as in in vitro grown oocyte-and-granulosa cell complexes.

    Science.gov (United States)

    Munakata, Yasuhisa; Kawahara-Miki, Ryoka; Shiratsuki, Shogo; Tasaki, Hidetaka; Itami, Nobuhiko; Shirasuna, Koumei; Kuwayama, Takehito; Iwata, Hisataka

    2016-08-25

    Follicle development is accompanied by proliferation of granulosa cells and increasing oocyte size. To obtain high-quality oocytes in vitro, it is important to understand the processes that occur in oocytes and granulosa cells during follicle development and the differences between in vivo and in vitro follicle development. In the present study, oocytes and granulosa cells were collected from early antral follicles (EAFs, 0.5-0.7 mm in diameter), small antral follicles (SAFs, 1-3 mm in diameter), large antral follicles (LAFs, 3-7 mm in diameter), and in vitro grown oocyte-and-granulosa cell complexes (OGCs), which were cultured for 14 days after collection from EAFs. Gene expression was analyzed comprehensively using the next-generation sequencing technology. We found top upstream regulators during the in vivo follicle development and compared them with those in in vitro developed OGCs. The comparison revealed that HIF1 is among the top regulators during both in vivo and in vitro development of OGCs. In addition, we found that HIF1-mediated upregulation of glycolysis in granulosa cells is important for the growth of OGCs, but the cellular metabolism differs between in vitro and in vivo grown OGCs. Furthermore, on the basis of comparison of upstream regulators between in vivo and in vitro development of OGCs, we believe that low expression levels of FLT1 (VEGFA receptor), SPP1, and PCSK6 can be considered causal factors of the suboptimal development under in vitro culture conditions.

  4. Research progress of preparation technology of lutein extract from marigold%制备万寿菊叶黄素技术研究进展

    Institute of Scientific and Technical Information of China (English)

    吴兴壮; 张华; 王小鹤; 鲁明; 张晓黎

    2012-01-01

    As the lutein is widespread in nature dihydroxy carotenoids, and it also is a human eye retina main component of macular pigment.Lutein could be an effective adjuvant therapy to prevent and age-related macular disease and cataract and other degenerative eye diseases,and its used in the field of bioactive substances have a wide application prospect.Marigold is the industrial extraction and separation on the ideal of industrial raw materials lutein.ln this paper, lutein extracted from marigold exclusive varieties,the molecular structure of lutein and lutein stability at home and abroad in recent years,six aspects of the relevant research work of marigold lutein and the prospect were reviewed.%由于叶黄素(1utein)是自然界广泛存在的二羟基类胡萝l、素,也是人眼视网膜黄斑色素主要组成部分。叶黄素可有效预防并辅助治疗老年性黄斑退化病和白内障等眼部疾病,其在生物活性物质利用领域有广泛的应用前景。万寿菊是工业上提取分离叶黄素的理想工业原料。本文从万寿菊叶黄素提取专用品种、叶黄素分子结构及叶黄素稳定性等6个方面综述了近年来国内外有关万寿菊叶黄素的研究工作,并进行了前景展望。

  5. Beverages formulated with whey protein and added lutein

    Directory of Open Access Journals (Sweden)

    Juliana de Cássia Gomes Rocha

    Full Text Available ABSTRACT: This study aimed to develop and characterize beverages formulated with whey protein and added lutein. Beverages formulated with 0.5 (F1, 2.0 (F2, 4.0 (F3 and 6.0% w/v (F4 whey protein were physicochemically and microbiologically characterized, and sensory evaluated. The physicochemical analyses indicated that the protein content significantly changed (P0.05 with increased protein content. The F2 formulation showed the highest sensory acceptance. Beverages offer a promising alternative to whey use and enhance the value of the product by the addition of lutein.

  6. The imitation switch ATPase Snf2l is required for superovulation and regulates Fgl2 in differentiating mouse granulosa cells.

    Science.gov (United States)

    Pépin, David; Paradis, François; Perez-Iratxeta, Carol; Picketts, David J; Vanderhyden, Barbara C

    2013-06-01

    Imitation switch (ISWI) proteins are catalytic subunits of chromatin remodeling complexes that alter nucleosome positioning by hydrolyzing ATP to regulate access to DNA. In mice, there are two paralogs, SNF2-homolog (SNF2H) and SNF2-like (SNF2L), which participate in different complexes and have contrasting patterns of expression. Here we investigate the role of SNF2L in ovaries by characterizing a mouse bearing an inactivating deletion of exon 6 that disrupts the ATPase domain. Snf2l mutant mice produce significantly fewer eggs than control mice when superovulated. Gonadotropin stimulation leads to a significant deficit in secondary follicles and an increase in abnormal antral follicles. Mutant females also failed to induce fibrinogen-like 2 (Fgl2) in response to human chorionic gonadotropin (hCG) stimulation, while overexpression of SNF2L was sufficient to drive its expression in granulosa cells. SNF2L was also shown to directly interact with the nuclear receptor co-activator flightless I (FLI-I) as shown by immunoprecipitation. These results begin to establish a role for SNF2L in the precise coordination of gene expression in granulosa cells during folliculogenesis and its broader implications in fertility.

  7. Lineage Specification of Ovarian Theca Cells Requires Multi-Cellular Interactions via Oocyte and Granulosa Cells

    Science.gov (United States)

    Liu, Chang; Peng, Jia; Matzuk, Martin M.; Yao, Humphrey H-C

    2015-01-01

    Organogenesis of the ovary is a highly orchestrated process involving multiple lineage determinations of ovarian surface epithelium, granulosa cells, and theca cells. While the sources of ovarian surface epithelium and granulosa cells are known, the origin(s) of theca progenitor cells have not been definitively identified. Here we show that theca cells derive from two sources: Wt1+ cells indigenous to the ovary and Gli1+ mesenchymal cells migrated from the mesonephros. These progenitors acquire theca lineage marker Gli1 in response to paracrine signals Desert hedgehog (Dhh) and Indian hedgehog (Ihh) from granulosa cells. Ovaries lacking Dhh/Ihh exhibit theca layer loss, blunted steroid production, arrested folliculogenesis, and failure to form corpora lutea. Production of Dhh/Ihh in granulosa cells requires Growth differentiation factor 9 (GDF9) from the oocyte. Our studies provide the first genetic evidence for the origins of theca cells and reveal a multicellular interaction critical for the formation of a functional theca. PMID:25917826

  8. Inhibition of NF-κB promotes autophagy via JNK signaling pathway in porcine granulosa cells

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    Gao, Hui; Lin, Lu; Haq, Ihtesham Ul; Zeng, Shen-ming, E-mail: zengshenming@gmail.com

    2016-04-22

    The transcription factor nuclear factor-κB (NF-κB) plays an important role in diverse processes, including cell proliferation and differentiation, apoptosis and inflammation. However, the role of NF-κB in porcine follicle development is not clearly elucidated. In this study, we demonstrated that follicle stimulating hormone (FSH) increased the level of inhibitor of NF-κB (IκB) protein and promoted the cytoplasmic localization of p65, indicating that FSH inhibits the activation of NF-κB in porcine granulosa cells. Moreover, inhibition of NF-κB by FSH or another specific inhibitor of NF-κB, pyrrolidine dithiocarbamate (PDTC), could activate JNK signaling and enhance autophagic activity in porcine granulosa cells. Knockdown of RelA (p65) Subunit of NF-κB by RNA interference abrogated the activation of JNK signaling pathway and the increase of autophagic protein expression by FSH. Meanwhile, the functional significance of FSH or PDTC-mediated autophagy were further investigated. Our results demonstrated that the increased autophagy promoted progesterone secretion in porcine granulosa cells. Blockage of autophagy by chloroquine obviated the FSH or PDTC-induced progesterone production. Taken together, these results indicate that inhibition of NF-κB increased autophagy via JNK signaling, and promote steroidogenesis in porcine granulosa cells. Our results provide new insights into the regulation and function of autophagy in mammalian follicle development. - Highlights: • FSH inhibits the activation of NF-κB in porcine primary granulosa cells. • Inhibition of NF-κB by FSH promotes autophagy via JNK signaling in granulosa cells. • Increased autophagy contributes to progesterone production in granulosa cells. • This is the first report against beclin1 regulation in porcine granulosa cells.

  9. Granulosa cell tumor of ovary: A clinicopathological study of four cases with brief review of literature

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    B R Vani

    2014-01-01

    Full Text Available Introduction: Adult granulosa cell tumor (GCT is a rare ovarian malignancy having good prognosis in comparison with other epithelial tumors. The study aims to collect data of all granulosa cell tumors diagnosed in ESIC Medical College & PGIMSR, Rajajinagar, Bangalore over the last 3 years and to describe the patient profile, ultrasonographic and various histopathological features.Materials and Methods: A total of 4 granulosa cell tumors were diagnosed in ESIC Medical College & PGIMSR, Rajajinagar, Bangalore during the period from June 2010 to June 2013. The patient′s age, clinical manifestations, radiological and histopathological findings were evaluated.Results: All 4 patients were diagnosed as adult granulosa cell tumor, three of four cases were in premenopausal age group and one case was in perimenopausal age. The clinical manifestations were menorrhagia and abdominal pain. Ultrasonographically, 2 cases of granulosa cell tumors were both solid and cystic and one case each was either solid or cystic. Histologically, variety of patterns like diffuse, trabecular, cords, spindle and clear cells were noted. Both Call-Exner bodies and nuclear grooves were observed in all cases. All four cases showed simple hyperplasia without atypia endometrial findings. Follow up on all patients revealed no evidence of recurrence.Conclusion: Granulosa cell tumor of the ovary is a rare ovarian entity. The important prognostic factor is staging of the tumor. Staging and histopathology helps in prediction of survival. Also diligent endometrial pathology has to be sorted to rule out endometrial carcinoma.

  10. Proprotein convertase furin regulates apoptosis and proliferation of granulosa cells in the rat ovary.

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    Xiaokui Yang

    Full Text Available Folliculogenesis is tightly controlled by a series of hormones, growth factors and cytokines, many of which are secreted as proproteins and require processing by proteases before becoming functional. Furin is a member of the subtilisin-like proteases that activate large numbers of proprotein substrates and is ubiquitously expressed and implicated in many physiological and pathological processes. However, the precise role of furin during folliculogenesis has not been thoroughly investigated. The goal of the present work is to identify the role of furin in the development of granulosa cells during folliculogenesis, using immunohistochemistry, RT-PCR, Western blot and functional studies in primary cultured rat granulosa cells. Our results demonstrate that furin is highly expressed in granulosa cells and oocytes of the ovary with very limited expression in other ovarian cells such as the epithelial, stromal or theca cells. Furin siRNA significantly increases apoptosis of the granulosa cells from large antral/preovulatory follicles, in part via downregulation of the anti-apoptotic proteins, XIAP and p-AKT. On the contrary, furin siRNA markedly decreases proliferation of granulosa cells based on the downregulation of proliferation cell nuclear antigen (PCNA. Taken together, these data suggest that furin may play an important role in regulating apoptosis and proliferation of granulosa cells.

  11. Effects of Fenvalerate on Steroidogenesis in Cultured Rat Granulosa Cells

    Institute of Scientific and Technical Information of China (English)

    JIAN-FENG CHEN; GUI-DONG DAI; XIN-RU WANG; HAI-YAN CHEN; RU LIU; JUN HE; LIN SONG; QIAN BIAN; LI-CHUN XU; JIAN-WEI ZHOU; HANG XIAO

    2005-01-01

    Objective This study was designed to examine the in vitro effects of fenvalerate on steroid production and steroidogenic enzymes mRNA expression level in rat granulosa cells. Methods Using primary cultured rat granulosa cells (rGCs) as model, fenvalerate of various concentrations (0, 1, 5, 25, 125, 625 μmol/L) was added to the medium for 24 h. In some cases, optimal concentrations of 22(R)-hydroxycholesterol (25 μmol/L), Follicle stimulating hormone (FSH, 2 mg/L), or 8-Bromo-cAMP (1 mmol/L) were provided. Concentrations of 17β-estradiol(E2) and progesterone (P4) in the medium from the same culture wells were measured by RIA and the steroidogenic enzyme mRNA level was quantified by semi-quantitative RT-PCR. Results Fenvalerate decreased both P4 and E2 production in a dose-dependent manner while it could significantly stimulate rGCs proliferation. This inhibition was stronger in the presence of FSH. Furthermore, it could not be reversed by 22(R)-hydroxycholesterol or 8-Bromo-cAMP. RT-PCR revealed that fenvalerate had no significant effect on 3β-HSD, but could increase the P450scc mRNA level. In addition, 17β-HSD mRNA level was dramatically reduced with the increase of fenvalerate dose after 24 h treatment. Conclusion Fenvalerate inhibits both P4 and E2 production in rGCs. These results support the view that fenvalerate is considered as a kind of endocrine-disrupting chemicals. The mechanism of its disruption may involve the effects on steroidogenesis signaling cascades and/or steroidogenic enzyme's activity.

  12. Multiple luteinizing hormone receptor (LHR protein variants, interspecies reactivity of anti-LHR mAb clone 3B5, subcellular localization of LHR in human placenta, pelvic floor and brain, and possible role for LHR in the development of abnormal pregnancy, pelvic floor disorders and Alzheimer's disease

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    Fernando Romaine I

    2003-06-01

    Full Text Available Abstract Distinct luteinizing hormone receptor (LHR protein variants exist due to the posttranslational modifications. Besides ovaries, LHR immunoreactivity (LHRI was also found in other tissues, such as the brain, fallopian tube, endometrium, trophoblast and resident tissue macrophages. The 3B5 mouse monoclonal antibody was raised against purified rat LHR. In rat, porcine and human ovaries, the 3B5 identified six distinct LHR bands migrating at ~92, 80, 68, 59, 52 and 48 kDa. Characteristic LHRI was detected in rat, human and porcine corpora lutea. During cellular differentiation, subcellular LHR distribution changed from none to granular cytoplasmic, perinuclear, surface, nuclear and no staining. There were also differences in vascular LHR expression – lack of LHRI in ovarian vessels and strong staining of vessels in other tissues investigated. In normal human term placentae, villous LHRI was associated with blood sinusoids and cytotrophoblast cells, and rarely detected in trophoblastic syncytium. In all abnormal placentae, the LHRI of sinusoids was absent, and syncytium showed either enhanced (immature placental phenotypes or no LHRI (aged placental phenotype. LHRI in human brain was identified in microglial cells (CD68+ resident macrophages. Protein extracts from human vaginal wall and levator ani muscle and fascia showed strong ~92 and 68 kDa species, and LHRI was detected in smooth muscle cells, fibroblasts, resident macrophages and nuclei of skeletal muscle fibers. Our observations indicate that, in contrast to the theory on the role of vascular hormone receptors in preferential pick up of circulating hormones, there is no need to enhance selective pick up rather only prevent LH/CG transport to inappropriate sites. Abnormal placental LHR expression may play a role in the development of abnormal pregnancy. Expression of LHR in the pelvic floor compartments suggests that high LH levels in postmenopausal women may contribute to the pelvic

  13. Multiple luteinizing hormone receptor (LHR) protein variants, interspecies reactivity of anti-LHR mAb clone 3B5, subcellular localization of LHR in human placenta, pelvic floor and brain, and possible role for LHR in the development of abnormal pregnancy, pelvic floor disorders and Alzheimer's disease.

    Science.gov (United States)

    Bukovsky, Antonin; Indrapichate, Korakod; Fujiwara, Hiroshi; Cekanova, Maria; Ayala, Maria E; Dominguez, Roberto; Caudle, Michael R; Wimalsena, Jay; Elder, Robert F; Copas, Pleas; Foster, James S; Fernando, Romaine I; Henley, Donald C; Upadhyaya, Nirmala B

    2003-06-03

    Distinct luteinizing hormone receptor (LHR) protein variants exist due to the posttranslational modifications. Besides ovaries, LHR immunoreactivity (LHRI) was also found in other tissues, such as the brain, fallopian tube, endometrium, trophoblast and resident tissue macrophages. The 3B5 mouse monoclonal antibody was raised against purified rat LHR. In rat, porcine and human ovaries, the 3B5 identified six distinct LHR bands migrating at approximately 92, 80, 68, 59, 52 and 48 kDa. Characteristic LHRI was detected in rat, human and porcine corpora lutea. During cellular differentiation, subcellular LHR distribution changed from none to granular cytoplasmic, perinuclear, surface, nuclear and no staining. There were also differences in vascular LHR expression--lack of LHRI in ovarian vessels and strong staining of vessels in other tissues investigated. In normal human term placentae, villous LHRI was associated with blood sinusoids and cytotrophoblast cells, and rarely detected in trophoblastic syncytium. In all abnormal placentae, the LHRI of sinusoids was absent, and syncytium showed either enhanced (immature placental phenotypes) or no LHRI (aged placental phenotype). LHRI in human brain was identified in microglial cells (CD68+ resident macrophages). Protein extracts from human vaginal wall and levator ani muscle and fascia showed strong approximately 92 and 68 kDa species, and LHRI was detected in smooth muscle cells, fibroblasts, resident macrophages and nuclei of skeletal muscle fibers. Our observations indicate that, in contrast to the theory on the role of vascular hormone receptors in preferential pick up of circulating hormones, there is no need to enhance selective pick up rather only prevent LH/CG transport to inappropriate sites. Abnormal placental LHR expression may play a role in the development of abnormal pregnancy. Expression of LHR in the pelvic floor compartments suggests that high LH levels in postmenopausal women may contribute to the pelvic

  14. Vitreoscilla hemoglobin gene ( vgb) improves lutein production in Chlorella vulgaris

    Science.gov (United States)

    Ma, Ruijuan; Lin, Xiangzhi

    2014-03-01

    Vitreoscilla hemoglobin is an oxygen-binding protein that promotes oxygen delivery and reduces oxygen consumption under low oxygen conditions to increase the efficiency of cell respiration and metabolism. In this study, we introduced a Vitreoscilla hemoglobin gene ( vgb) into Chlorella vulgaris by Agrobacterium tumefaciens -mediated transformation (ATMT). PCR analysis confirmed that the vgb gene was successfully integrated into the Chlorella vulgaris genome. Analysis of biomass obtained in shake flasks revealed transformant biomass concentrations as high as 3.28 g/L, which was 38.81% higher than that of the wild-type strain. Lutein content of transformants also increased slightly. Further experiments recovered a maximum lutein yield of 2.91 mg/L from the transformants, which was 36.77% higher than that of the wild-type strain. The above results suggest that integrated expression of the vgb gene may improve cell growth and lutein yield in Chlorella vulgaris, with applications to lutein production from Chlorella during fermentation.

  15. Biochemical System Analysis of Lutein Production by Heterotrophic Chlorella pyrenoidosa in a Fermentor

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    Zheng-Yun Wu

    2009-01-01

    Full Text Available Chlorella is a promising alternative source of lutein, as it can be cultivated heterotrophically with high efficiency. In this study, the carotenoids in Chlorella pyrenoidosa heterotrophically cultivated in a 19-litre fermentor have been analyzed and determined by using HPLC and HPLC-MS. A biochemical system theory (BST model was developed for understanding the regulatory features of carotenoid metabolism during the batch cultivation. Factors that influence lutein production by C. pyrenoidosa were discussed based on the model. It shows that low flux for lycopene formation is the major bottleneck for lutein production, while by-product syntheses and inhibitions affect the cellular lutein content much less. However, with further increase of the cellular lutein content, the inhibition on lycopene formation by lutein may become a limiting factor. Although speculative, these results may provide useful information for further elucidation of the regulatory mechanisms of carotenoid biosynthesis in Chlorella and modifying its metabolic network to enhance lutein production.

  16. Isolation and purification of lutein from the microalga Chlorella vulgaris by extraction after saponification.

    Science.gov (United States)

    Li, Hua-Bin; Jiang, Yue; Chen, Feng

    2002-02-27

    A simple and efficient method for the isolation and purification of lutein from the microalga Chlorella vulgaris was developed. Crude lutein was obtained by extraction with dichloromethane from the microalga after saponification. Partition values of lutein in the two-phase system of ethanol-water-dichloromethane at different ratios were measured by HPLC so as to assist the determination of an appropriate condition for washing water-soluble impurities in the crude lutein. Partition values of lutein in another two-phase system of ethanol-water-hexane at different ratios were also measured by HPLC for determining the condition for removing fat-soluble impurities. The water-soluble impurities in the crude lutein were removed by washing with 30% aqueous ethanol, and the fat-soluble impurities were removed by extraction with hexane. The final purity of lutein obtained was 90-98%, and the yield was 85-91%.

  17. The stimulatory effect of albumin on luteinizing hormone-stimulated Leydig cell steroid production depends on its fatty acid content and correlates with conformational changes

    NARCIS (Netherlands)

    R. Melsert (R.); O.J.M. Bos (O. J M); R.F. van der Linden (R.); M.J.E. Fischer (M. J E); S.M. Wilting (Saskia); L.H.M. Janssen (Lambert); J.W. Hoogerbrugge (Jos); F.F.G. Rommerts (Focko)

    1991-01-01

    markdownabstract__Abstract__ The effects of purified albumin species and albumin fragments (0.2–1% w/v) on short-term (4 h) steroid secretion by immature rat Leydig cells, in the presence of a maximally stimulating dose of luteinizing hormone (LH), were investigated. Human albumin and the peptic fr

  18. Lutein Induces Autophagy via Beclin-1 Upregulation in IEC-6 Rat Intestinal Epithelial Cells.

    Science.gov (United States)

    Chang, Chi-Jen; Lin, Ji-Fan; Hsiao, Chien-Yu; Chang, Hsun-Hao; Li, Hsin-Ju; Chang, Hsun-Hsien; Lee, Gon-Ann; Hung, Chi-Feng

    2017-01-01

    Lutein is a carotenoid with anti-oxidant properties. Autophagy, an evolutionarily conserved catabolic cellular pathway for coping with stress conditions, is responsive to reactive oxygen species (ROS) and degrades damaged organelles. We previously demonstrated that lutein can induce anti-oxidant enzymes to relieve methotrexate-induced ROS stress. We therefore hypothesized that lutein, which activates ROS-scavenging enzymes, can also induce autophagy for cell survival. In this study, we demonstrated that lutein treatment attenuated the reduction in cell viability caused by H2O2. Lutein dose-dependently induced the processing of microtubule-associated protein light chain 3 (LC3)-II, an autophagy marker protein, and accumulation of LC3-positive puncta in rat intestinal IEC-6 cells. Furthermore, (a) direct observation of autophagosome formation through transmission electron microscopy, (b) upregulation of autophagy-related genes including ATG4A, ATG5, ATG7, ATG12, and beclin-1 (BENC1), and (c) increased BECN1/Bcl-2 ratio confirmed the induction of autophagy by lutein. The results revealed that bafilomycin-A1-induced inhibition of autophagy reduced cell viability and increased apoptosis in lutein-treated cells, indicating a protective role of lutein-induced autophagy. Lutein treatment also activated adenosine monophosphate-activated protein kinase (AMPK), c-Jun N-terminal kinase (JNK), and p-38, but had no effects on the induction of extracellular signal-related kinase or inhibition of mTOR; however, the inhibition of activated AMPK, JNK, or p-38 did not attenuate lutein-induced autophagy. Finally, increased BECN1 expression levels were detected in lutein-treated cells, and BECN1 knockdown abolished autophagy induction. These results suggest that lutein-induced autophagy was mediated by the upregulation of BECN1 in IEC-6 cells. We are the first to demonstrate that lutein induces autophagy. Elevated autophagy in lutein-treated IEC-6 cells may have a protective role

  19. A regulatory role of androgen in ovarian steroidogenesis by rat granulosa cells.

    Science.gov (United States)

    Hasegawa, Toru; Kamada, Yasuhiko; Hosoya, Takeshi; Fujita, Shiho; Nishiyama, Yuki; Iwata, Nahoko; Hiramatsu, Yuji; Otsuka, Fumio

    2017-09-01

    Excess androgen and insulin-like growth factor (IGF)-I in the ovarian follicle has been suggested to be involved in the pathophysiology of polycystic ovary syndrome (PCOS). Here we investigated the impact of androgen and IGF-I on the regulatory mechanism of ovarian steroidogenesis using rat primary granulosa cells. It was revealed that androgen treatment with dihydrotestosterone (DHT) amplified progesterone synthesis in the presence of FSH and IGF-I, whereas it had no significant effect on estrogen synthesis by rat granulosa cells. In accordance with the effects of androgen on steroidogenesis, DHT enhanced the expression of progesterogenic factors and enzymes, including StAR, P450scc and 3βHSD, and cellular cAMP synthesis induced by FSH and IGF-I. Of note, treatment with DHT and IGF-I suppressed Smad1/5/8 phosphorylation and transcription of the BMP target gene Id-1, suggesting that androgen and IGF-I counteract BMP signaling that inhibits FSH-induced progesterone synthesis in rat granulosa cells. DHT was revealed to suppress the expression of BMP-6 receptors, consisting of ALK-2, ALK-6 and ActRII, while it increased the expression of inhibitory Smads in rat granulosa cells. In addition, IGF-I treatment upregulated androgen receptor (AR) expression and DHT treatment suppressed IGF-I receptor expression on rat granulosa cells. Collectively, the results indicate that androgen and IGF-I mutually interact and accelerate progesterone production, at least in part, by regulating endogenous BMP signaling in rat granulosa cells. Cooperative effects of androgen and IGF-I counteract endogenous BMP-6 activity in rat granulosa cells, which is likely to be functionally linked to the steroidogenic property shown in the PCOS ovary. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Estrogen receptor immunoreactivity is present in the majority of central histaminergic neurons: evidence for a new neuroendocrine pathway associated with luteinizing hormone-releasing hormone-synthesizing neurons in rats and humans.

    Science.gov (United States)

    Fekete, C S; Strutton, P H; Cagampang, F R; Hrabovszky, E; Kalló, I; Shughrue, P J; Dobó, E; Mihály, E; Baranyi, L; Okada, H; Panula, P; Merchenthaler, I; Coen, C W; Liposits, Z S

    1999-09-01

    The central regulation of the preovulatory LH surge requires a complex sequence of interactions between neuronal systems that impinge on LH-releasing hormone (LHRH)-synthesizing neurons. The reported absence of estrogen receptors (ERs) in LHRH neurons indicates that estrogen-receptive neurons that are afferent to LHRH neurons are involved in mediating the effects of this steroid. We now present evidence indicating that central histaminergic neurons, exclusively located in the tuberomammillary complex of the caudal diencephalon, serve as an important relay in this system. Evaluation of this system revealed that 76% of histamine-synthesising neurons display ERalpha-immunoreactivity in their nucleus; furthermore histaminergic axons exhibit axo-dendritic and axo-somatic appositions onto LHRH neurons in both the rodent and the human brain. Our in vivo studies show that the intracerebroventricular administration of the histamine-1 (H1) receptor antagonist, mepyramine, but not the H2 receptor antagonist, ranitidine, can block the LH surge in ovariectomized estrogen-treated rats. These data are consistent with the hypothesis that the positive feedback effect of estrogen in the induction of the LH surge involves estrogen-receptive histamine-containing neurons in the tuberomammillary nucleus that relay the steroid signal to LHRH neurons via H1 receptors.

  1. In vitro changes in porcine ovarian granulosa cells induced by copper.

    Science.gov (United States)

    Roychoudhury, Shubhadeep; Bulla, Jozef; Sirotkin, Alexander V; Kolesarova, Adriana

    2014-01-01

    Objective of this in vitro study was to examine the secretion activity (progesterone and insulin-like growth factor I) of porcine ovarian granulosa cells after copper (Cu) addition and to outline a potential intracellular mediator (cyclin B1) of its effects. It also aimed at investigating the apoptotic potential of Cu on porcine ovarian granulosa cells after addition in vitro. Ovarian granulosa cells were incubated with copper sulphate (CuSO4·5H2O) at the doses 0.33, 0.40, 0.50, 1.0 and 2.0 μL mL(-1) for 18 h and compared with control group without Cu addition. Release of progesterone (P4) and insulin-like growth factor I (IGF-I) by granulosa cells was assessed by RIA, expression of cyclin B1 by immunocytochemistry and apoptosis by TUNEL assay. Observations show that P4 release by granulosa cells was inhibited while the release of IGF-I and cyclin B1 was stimulated significantly (P < 0.05) by CuSO4·5H2O addition at the dose 2.0 μL mL(-1). Also, addition of CuSO4.5H2O at the lowest dose used in the study (0.33 μL mL(-1)) significantly (P < 0.05) decreased apoptosis in granulosa cells. In conclusion, results indicate dose dependent effect of Cu on (1) secretion of steroid hormone progesterone and growth factor IGF-I, (2) expression of cyclin B1 as marker of proliferation of porcine ovarian granulosa cells, (3) apoptosis of porcine ovarian granulosa cells and, (4) that the effect of Cu on ovarian cell proliferation could be mediated by IGF-I and cyclin B1. Obtained data suggest interference of Cu in the pathways of proliferation of porcine ovarian granulosa cells through hormonal and intracellular peptide cyclin B1.

  2. MC-LR Exposure Leads to Subfertility of Female Mice and Induces Oxidative Stress in Granulosa Cells

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    Jiang Wu

    2015-12-01

    Full Text Available Health risk of human exposure to microcystin-leucine arginine (MC-LR has aroused more and more attention over the past few decades. In the present study, MC-LR was orally administered to female mice at 0, 1, 10 and 40 μg/L for three and six months. We found that chronic exposure to MC-LR at environmental levels could stimulate follicle atresia and lead to decreased developmental follicles, accompanied by a reduction of gonadosomatic index (GSI. In line with the irregular gonadal hormone level and estrus cycles, subfertility of female mice was also confirmed by analyzing numbers of litters and pups. The in vitro study suggested that granulosa cells could uptake MC-LR and should be the target of the toxicant. Oxidative stress in granulose cells induced by MC-LR promoted follicle atresia and eventually leads to female subfertility.

  3. Hemoglobin: a gas transport molecule that is hormonally regulated in the ovarian follicle in mice and humans.

    Science.gov (United States)

    Brown, Hannah M; Anastasi, Marie R; Frank, Laura A; Kind, Karen L; Richani, Dulama; Robker, Rebecca L; Russell, Darryl L; Gilchrist, Robert B; Thompson, Jeremy G

    2015-01-01

    An increasing number of nonerythroid tissues are found to express hemoglobin mRNA and protein. Hemoglobin is a well-described gas transport molecule, especially for O2, but also for NO, CO2, and CO, and also acts as a reactive oxygen species scavenger. We previously found Hba-a1 and Hbb mRNA and protein at high levels within mouse periovulatory cumulus cells, but not in cumulus following in vitro maturation. This led us to investigate the temporal and spatial regulation in follicular cells during the periovulatory period. Cumulus-oocyte complexes were collected from equine chorionic gonadotropin/human chorionic gonadotropin-treated peripubertal SV129 female mice and collected and analyzed for gene expression and protein localization at a variety of time points over the periovulatory period. A further cohort matured in vitro with different forms of hemoglobin (ferro- and ferrihemoglobin) under different O2 atmospheric conditions (2%, 5%, and 20% O2) were subsequently fertilized in vitro and cultured to the blastocyst stage. Murine mRNA transcripts for hemoglobin were regulated by stimulation of the ovulatory cascade, in both granulosa and cumulus cells, and expression of HBA1 and HBB was highly significant in human granulosa and cumulus, but erythrocyte cell marker genes were not. Several other genes involved in hemoglobin function were similarly luteinizing hormone-regulated, including genes for heme biosynthesis. Immunohistochemistry revealed a changing localization pattern of HBA-A1 protein in murine cumulus cells and oocytes following the ovulatory signal. Significantly, no positive staining for HBA-A1 protein was observed within in vitro-matured oocytes, but, if coincubated with ferro- or ferrihemoglobin, cytoplasmic HBA-A1 was observed, similar to in vivo-derived oocytes. Addition of ferro-, but not ferrihemoglobin, had a small, positive effect on blastocyst yield, but only under either 2% or 20% O2 gas atmosphere. The identification of hemoglobin within

  4. Granulosa cell tumor of the ovary and antecedent of adjuvant tamoxifen use for breast cancer

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    Abahssain Halima

    2010-08-01

    Full Text Available Abstract Background Adult granulosa cell tumor associated with antecedent use of tamoxifen as adjuvant hormonotherapy for breast cancer is rare. The pathogenesis of this occurrence remains difficult to explain. The estrogenic effect of tamoxifen can be one such explanation. Case presentation A 47 year-old women was treated with surgery, chemotherapy, radiotherapy and tamoxifen for stage III estrogen receptor positive breast carcinoma. Ten months after stopping tamoxifen, we diagnosed a stage Ic granulosa cell tumor of the ovary. Conclusions Use of tamoxifen has been found to be associated with gynecological tumors like endometrial carcinoma. Its association with granulosa cell tumor of the ovary is uncommon. Only two previous cases have been reported in literature.

  5. Determination of Lutein from Fruit and Vegetables Through an Alkaline Hydrolysis Extraction Method and HPLC Analysis.

    Science.gov (United States)

    Fratianni, Alessandra; Mignogna, Rossella; Niro, Serena; Panfili, Gianfranco

    2015-12-01

    A simple and rapid analytical method for the determination of lutein content, successfully used for cereal matrices, was evaluated in fruit and vegetables. The method involved the determination of lutein after an alkaline hydrolysis of the sample matrix, followed by extraction with solvents and analysis by normal phase HPLC. The optimized method was simple, precise, and accurate and it was characterized by few steps that could prevent loss of lutein and its degradation. The optimized method was used to evaluate the lutein amounts in several fruit and vegetables. Rich sources of lutein were confirmed to be green vegetables such as parsley, spinach, chicory, chard, broccoli, courgette, and peas, even if in a range of variability. Taking into account the suggested reference values these vegetables can be stated as good sources of lutein. © 2015 Institute of Food Technologists®

  6. Spatial differences within the membrana granulosa in the expression of focimatrix and steroidogenic capacity.

    Science.gov (United States)

    Nguyen, Tracy; Lee, Samuel; Hatzirodos, Nicholas; Hummitzsch, Katja; Sullivan, Thomas R; Rodgers, Raymond J; Irving-Rodgers, Helen F

    2012-11-05

    In the ovarian follicular membrana granulosa there are morphological and functional differences between cells adjacent to the follicular fluid lumen, or aligning the basal lamina. Amongst the observed functional differences are steroidogenic capacity and expression levels of a novel basal lamina, focimatrix; both of which increase in the later stages of antral follicle growth. A number of different studies have produced apparently inconsistent results as to which cell layers are more steroidogenic. To examine this systematically, individual bovine follicles, confirmed as healthy by post hoc histological examination, were used to isolate populations of apical and basal granulosa cells. Cell counts revealed that the respective groups did not differ in the numbers of cells, thus confirming the separation of these populations. We measured gene expression (quantitative RT-PCR, n=8-10, follicle diameter 14.0±0.5 mm) and protein levels (Western immunoblotting, n=14, follicle diameter 11.9±0.5 mm) and hormone production from granulosa cells (2.5×10(5) viable cells/well in serum-free conditions for 24 h, n=15, diameter 12±0.5 mm). Levels of mRNA of HSD3B1 and CYP19A1 and three focimatrix genes COL4A1, HSPG2 and LAMB2 and LHCGR were significantly lower in apical granulosa cells (P0.05). The protein levels of steroidogenic enzymes P450scc and P450arom were significantly higher in apical cells (P0.05). Progesterone production was significantly lower and oestradiol production was significantly higher in apical granulosa cells (Pmembrana granulosa. Discrepancies in the literature on their steroidogenic capacity may reflect differences in the steroidogenic parameters measured. Crown Copyright © 2012. Published by Elsevier Ireland Ltd. All rights reserved.

  7. Effects of transferrin on aromatase activity in porcine granulosa cells in vitro.

    Directory of Open Access Journals (Sweden)

    Małgorzata Duda

    2009-01-01

    Full Text Available Proliferating cells have an absolute requirement for iron, which is delivered by transferrin with subsequent intracellular transport via the transferrin receptor. Recent studies have reported that transferrin plays a crucial role in the local regulation of ovarian function, apart from its iron-binding characteristic. Therefore, the present study was undertaken to explore the possible role of transferrin in porcine granulosa cells function by examining its influence on aromatase activity, the most important indicator of follicular cell differentiation. In the first series of studies, pig granulosa cells isolated from small, immature follicles were cultured in the presence of transferrin alone (10 microg/ml or 100 microg/ml or with the addition of FSH (100ng/ml. The second series of studies was undertaken to determine transferrin-stimulated granulosa cells ability to aromatize exogenous testosterone (1x10(-7M. One hour after the establishment of cultures an aromatase inhibitor CGS16949A was added to test its influence on estradiol production. After 48 hours, cultures were terminated and cells were processed for immunocytochemical staining of aromatase. Media were frozen for further estradiol level analysis. Positive immunostaining for aromatase was found in all granulosa cell cultures. The intensity of immunostaining was always stronger in cultures supplemented with FSH whereas the addition of transferrin had no effect. Granulosa cells in vitro synthesized the highest amount of estradiol after the addition of FSH and exogenous testosterone as measured radioimmunologically. Concomitant treatment with FSH and transferrin caused an inhibition of FSH-stimulated aromatase activity. The production of estradiol also declined in the presence of FSH, testosterone and transferrin. This study demonstrates that transferrin had a dose-dependent inhibitory effect on FSH-stimulated aromatase activity, which was confirmed by radioimmunoassay. Our results indicate

  8. Melatonin modulates the functions of porcine granulosa cells via its membrane receptor MT2 in vitro.

    Science.gov (United States)

    He, Ya-Mei; Deng, Hong-Hui; Shi, Mei-Hong; Bodinga, Bello Musa; Chen, Hua-Li; Han, Zeng-Sheng; Jiang, Zhong-Liang; Li, Qing-Wang

    2016-09-01

    Melatonin (N-acetyl-5-methoxytryptamine) is documented as a hormone involved in the circadian regulation of physiological and neuroendocrine function in mammals. Herein, the effects of melatonin on the functions of porcine granulosa cells in vitro were investigated. Porcine granulosa cells were cultivated with variable concentrations of melatonin (0, 0.001, 0.01, 0.1, 1.0, and 10ng/mL) for 48h. Melatonin receptor agonist (IIK7) and antagonist (Luzindole, 4P-PDOT) were used to further examine the action of melatonin. The results showed optimum cell viability and colony-forming efficiency of porcine granulosa cells at 0.01ng/mL melatonin for 48-h incubation period. The percentage of apoptotic granulosa cells was significantly reduced by 0.01 and 0.1ng/mL melatonin within the 48-h incubation period as compared with the rest of the treatments. Estradiol biosynthesis was significantly stimulated by melatonin supplementation and suppressed for the progesterone secretion; the minimum ratio of progesterone to estradiol was 1.82 in 0.01ng/mL melatonin treatment after 48h of cultivation. Moreover, the expression of BCL-2, CYP17A1, CYP19A1, SOD1, and GPX4 were up-regulated by 0.01ng/mL melatonin or combined with IIK7, but decreased for the mRNA levels of BAX, P53, and CASPASE-3, as compared with control or groups treated with Luzindole or 4P-PDOT in the presence of melatonin. In conclusion, the study demonstrated that melatonin mediated proliferation, apoptosis, and steroidogenesis in porcine granulosa cells predominantly through the activation of melatonin receptor MT2 in vitro, which provided evidence of the beneficial role of melatonin as well as its functional mechanism in porcine granulosa cells in vitro.

  9. Effect of the methoxychlor metabolite HPTE on the rat ovarian granulosa cell transcriptome in vitro.

    Science.gov (United States)

    Harvey, Craig N; Esmail, Mahmoud; Wang, Qi; Brooks, Andrew I; Zachow, Rob; Uzumcu, Mehmet

    2009-07-01

    Ovarian granulosa cells play a central role in steroidogenesis, which is critical for female reproduction. Follicle-stimulating hormone (FSH) promotes cyclic adenosine monophosphate (cAMP)-mediated signaling to regulate granulosa cell steroidogenesis. We have shown previously that 2,2-bis-(p-hydroxyphenyl)-1,1,1-trichloroethane (HPTE) inhibits FSH- and dibutyryl cAMP-stimulated steroidogenesis and affects the messenger RNA levels of steroidogenic pathway enzymes in rat granulosa cells. However, HPTE showed a differential effect in FSH- and cAMP-stimulated cells in that HPTE more completely blocked FSH- when compared to cAMP-driven steroidogenesis. The objective of this study was to analyze the effects of HPTE on global gene expression profiles in untreated granulosa cells and those challenged with FSH or cAMP. Granulosa cells from immature rats were cultured with 0, 1, 5, or 10 microM HPTE in the presence or absence of either 3 ng FSH/ml or 1mM cAMP for 48 h. Total RNA was isolated for real-time quantitative PCR and microarray analysis using the GeneChip Rat Genome 230 2.0 and ArrayAssist Microarray Suite. An investigation of changes in gene expression across all HPTE treatments showed that HPTE altered more genes in FSH- (approximately 670 genes) than in cAMP-stimulated cells (approximately 366 genes). Analysis confirmed that HPTE more effectively inhibited FSH- than cAMP-induced steroid pathway gene expression and steroidogenesis. Furthermore, expression patterns of novel genes regulating signal transduction, transport, cell cycle, adhesion, differentiation, motility and growth, apoptosis, development, and metabolism were all altered by HPTE. This study further established that HPTE exerts differential effects within the granulosa cell steroidogenic pathway and revealed that these effects include broader changes in gene expression.

  10. Management of Ocular Diseases Using Lutein and Zeaxanthin: What Have We Learned from Experimental Animal Studies?

    OpenAIRE

    Chunyan Xue; Richard Rosen; Adrienne Jordan; Dan-Ning Hu

    2015-01-01

    Zeaxanthin and lutein are two carotenoid pigments that concentrated in the retina, especially in the macula. The effects of lutein and zeaxanthin on the prevention and treatment of various eye diseases, including age-related macular degeneration, diabetic retinopathy and cataract, ischemic/hypoxia induced retinopathy, light damage of the retina, retinitis pigmentosa, retinal detachment, and uveitis, have been studied in different experimental animal models. In these animal models, lutein and ...

  11. Study on connexin gene and protein expression and cellular distribution in relation to real-time proliferation of porcine granulosa cells.

    Science.gov (United States)

    Kempisty, B; Ziółkowska, A; Ciesiółka, S; Piotrowska, H; Antosik, P; Bukowska, D; Nowicki, M; Brüssow, K P; Zabel, M

    2014-01-01

    Granulosa cells (GCs) play an important role during follicle growth and development in preovulatory stage. Moreover, the proteins such as connexins are responsible for formation of protein channel between follicular-cumulus cells and oocyte. This study was aimed to investigate the role of connexin expression in porcine GCs in relation to their cellular distribution and real-time cell proliferation. In the present study, porcine GCs were isolated from the follicles of puberal gilts and then cultured in a real-time cellular analyzer (RTCA) system for 168 h. The expression levels of connexins (Cxs) Cx36, Cx37, Cx40 and Cx43 mRNA were measured by RQ-PCR analysis, and differences in the expression and distribution of Cx30, Cx31, Cx37, Cx43 and Cx45 proteins were analyzed by confocal microscopic visualization. We found higher level of Cx36, Cx37, and Cx43 mRNA expression in GCs at recovery (at 0 h of in vitro culture, IVC) compared to all analyzed time periods of IVC (24, 48, 72, 96, 120, 144 and 168 h; Pproteins were higher before (0 h) compared to after 168 h of IVC. The expression of Cx30 and Cx43, however, did not vary between the groups. In all, the proteins were distributed throughout the cell membrane rather than in the cytoplasm both before and after IVC. After 24 h of IVC, we observed a significant increase in the proliferation of GCs (log phase). We found differences in the proliferation index between 72-96 and 96- 140 h within the same population of GCs. In conclusion, the decrease in the expression of Cx mRNAs and proteins following IVC could be associated with a breakdown in gap-junction connections (GJCs), and leads to the decreased of their activity, which may be a reason of non-functional existence of connexon in follicular granulosa cells. These data indicated that the differentiation and proliferation of GCs and lutein cells are regulated by distinct mechanisms in pigs.

  12. Granulosa Cell Tumor Juvenile - Case Report-Primary Care Tumor de Celulas da Granulosa Juvenil - Relato de Caso em Atenção Primária

    Directory of Open Access Journals (Sweden)

    Anelise Olmos Grings

    2010-11-01

    Full Text Available

    The authors given an account of children of 2 years and 4 months with ginaecology complaints (white vaginal secretion, diffuse abdominal pain and early pseudo-puberty. Beginning investigation in Unity of Health Care PSF of Porto Alegre, are refer to the secondary levei was diagnosis ovarian tumor (Juvenile Granulosa Cell Tumor Ovary.

    Os autores relatam o caso de urna criança de 2 anos e 4 meses com queixas ginecológicas (secreção vaginal branca, dor abdominal difusa e pseudopuberdade precoce (telarca. Iniciada investigação em Unidade de Saúde PSF de Porto Alegre, sendo referenciada ao nível secundário onde foi diagnosticada neoplasia ovariana (Tumor de Células da Granulosa Juvenil.

  13. Effect of Lutein and Antioxidant Supplementation on VEGF Expression, MMP-2 Activity, and Ultrastructural Alterations in Apolipoprotein E-Deficient Mouse

    Directory of Open Access Journals (Sweden)

    Patricia Fernández-Robredo

    2013-01-01

    Full Text Available Oxidative stress is involved in the pathogenesis of several diseases such as atherosclerosis and age-related macular degeneration (AMD. ApoE-deficient mice (apoE−/− are a well-established model of genetic hypercholesterolemia and develop retinal alterations similar to those found in humans with AMD. Thus supplementation with lutein or multivitamin plus lutein and glutathione complex (MV could prevent the onset of these alterations. ApoE−/− mice (n=40, 3 months old were treated daily for 3 months with lutein (AE-LUT or MV (two doses: AE-MV15 (15 mg/kg/day and AE-MV50 (50 mg/kg/day and were compared to controls with vehicle (AE-C. Wild-type mice (n=10 were also used as control (WT-C. ApoE−/− mice showed higher retinal lipid peroxidation and increased VEGF expression and MMP-2 activity, associated with ultrastructural alterations such as basal laminar deposits, vacuoles, and an increase in Bruch's membrane thickness. While lutein alone partially prevented the alterations observed in apoE−/− mice, MV treatment substantially reduced VEGF levels and MMP-2 activity and ameliorated the retinal morphological alterations. These results suggest that oxidative stress in addition to an increased expression and activity of proangiogenic factors could participate in the onset or development of retinal alterations of apoE−/− mice. Moreover, these changes could be prevented by efficient antioxidant treatments.

  14. Cloning and Expression of Luteinizing Hormone Subunits in Chinese Hamster Ovary Cell Line

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    Zeinab Soleimanifar

    2016-10-01

    Full Text Available Background: Luteinizing hormone (LH was secreted by the stimulating cells of the testes and ovaries in the anterior pituitary gland. The application of this hormone is in the treatment of men and women with infertility and amenorrhea respectively.Materials and Methods: In the present study the alpha and beta subunits of human LH gene were cloned into the pEGFP-N1 expression vector and produced the recombinant LH hormone in Chinese hamster ovary (CHO eukaryotic system.Results: Alpha and beta subunits of LH hormone were cloned between NheI and BamHI cut sites of pEGFP_N1 expression plasmid and confirmed by PCR.  Hormone expression was evaluated in CHO cell line by Western blotting using the specific antibody.Conclusion: Alpha and beta subunits of LH hormone were expressed in CHO cell line perfectly.

  15. Incidence of apoptotic bodies in membrana granulosa of the patients participating in an in vitro fertilization program.

    Science.gov (United States)

    Nakahara, K; Saito, H; Saito, T; Ito, M; Ohta, N; Sakai, N; Tezuka, N; Hiroi, M; Watanabe, H

    1997-02-01

    To investigate the incidence of apoptotic bodies in mural granulosa cell masses and cumulus cell masses. Nonrandomized, prospective study. Department of Obstetrics and Gynecology, Yamagata University School of Medicine, Yamagata, Japan. One hundred twenty-nine normally ovulating women underwent ovulation induction for IVF-ET with GnRH analogue (GnRH-a) and gonadotropins. Patients underwent follicle aspiration after the administration of hCG. The nuclei of recovered granulosa cells were examined by fluorescence microscopy and the incidence of apoptotic bodies was tabulated. The incidence of apoptotic bodies was significantly higher in mural granulosa cell masses than in cumulus cell masses in the entire group of 129 patients. Both incidence of apoptotic bodies of mural granulosa cell masses and cumulus cell masses were significantly higher in patients with less than six follicular oocytes compared with patients with six or more oocytes. Nonpregnant patients showed significantly higher incidence of apoptotic bodies in mural granulosa cell masses compared with pregnant patients. These results indicate that mural granulosa cell masses and cumulus cell masses may have different functions in follicular maturation. The incidence of apoptotic bodies in mural granulosa cell masses can be used as an indicator of success of IVF.

  16. Effect of vitamin D3 on production of progesterone in porcine granulosa cells by regulation of steroidogenic enzymes.

    Science.gov (United States)

    Hong, So-Hye; Lee, Jae-Eon; Kim, Hong Sung; Jung, Young-Jin; Hwang, DaeYoun; Lee, Jae Ho; Yang, Seung Yun; Kim, Seung-Chul; Cho, Seong-Keun; An, Beum-Soo

    2016-05-01

    1,25-dihydroxyvitamin D3 (VD3), an active form of Vitamin D, is photosynthesized in the skin of vertebrates in response to solar ultraviolet B radiation (UV-B). VD3 deficiency can cause health problems such as immune disease, metabolic disease, and bone disorders. It has also been demonstrated that VD3 is involved in reproductive functions. Female sex hormones such as estrogen and progesterone are biosynthesized mainly in ovarian granulosa cells as the ovarian follicle develops. The functions of sex hormones include regulation of the estrus cycle and puberty as well as maintenance of pregnancy in females. In this study, we isolated granulosa cells from porcine ovaries and cultured them for experiments. To examine the effects of VD3 on ovarian granulosa cells, the mRNA and protein levels of genes were analyzed by Real-time PCR and Western blotting assay. Production of progesterone from granulosa cells was also measured by ELISA assay. As a result, transcriptional and translational regulation of progesterone biosynthesis-related genes in granulosa cells was significantly altered by VD3. Furthermore, progesterone concentrations in porcine granulosa cell-cultured media decreased in response to VD3. These results show that VD3 was a strong regulator of sex steroid hormone production in porcine granulosa cells, suggesting that vitamin D deficiency may result in inappropriate sexual development of industrial animals and eventually economic loss.

  17. Pilidiella tibouchinae sp. nov. associated with foliage blight of Tibouchina granulosa (quaresmeira) in Brazil

    NARCIS (Netherlands)

    Miranda, B.E.; Barreto, R.W.; Crous, P.W.; Groenewald, J.Z.

    2012-01-01

    Tibouchina granulosa (Melastomataceae), Brazilian glorytree (Brazilian common name - quaresmeira), a common tree of the Atlantic Forest of Brazil, is widely used as an ornamental for its violet or pink blossoms. Little is known about fungal diseases affecting this species, although these represent a

  18. Bilateral occurrence of granulosa-theca cell tumors in an Arabian mare.

    Science.gov (United States)

    Frederico, Lisa M; Gerard, Mathew P; Pinto, Carlos R F; Gradil, Carlos M

    2007-05-01

    An Arabian mare was referred for right granulosa-theca cell tumor (GTCT) evaluation. The mare was presented 4.5 years later for a left GTCT, after successfully conceiving and delivering a normal foal in the interim. The concurrent or nonconcurrent occurrence of bilateral GTCT in mares appears to be rare.

  19. Anti-Mullerian hormone recruits BMPR-IA in immature granulosa cells.

    Directory of Open Access Journals (Sweden)

    Lauriane Sèdes

    Full Text Available Anti-Müllerian hormone (AMH is a member of the TGF-β superfamily secreted by the gonads of both sexes. This hormone is primarily known for its role in the regression of the Müllerian ducts in male fetuses. In females, AMH is expressed in granulosa cells of developing follicles. Like other members of the TGF-β superfamily, AMH transduces its signal through two transmembrane serine/threonine kinase receptors including a well characterized type II receptor, AMHR-II. The complete signalling pathway of AMH involving Smads proteins and the type I receptor is well known in the Müllerian duct and in Sertoli and Leydig cells but not in granulosa cells. In addition, few AMH target genes have been identified in these cells. Finally, while several co-receptors have been reported for members of the TGF-β superfamily, none have been described for AMH. Here, we have shown that none of the Bone Morphogenetic Proteins (BMPs co-receptors, Repulsive guidance molecules (RGMs, were essential for AMH signalling. We also demonstrated that the main Smad proteins used by AMH in granulosa cells were Smad 1 and Smad 5. Like for the other AMH target cells, the most important type I receptor for AMH in these cells was BMPR-IA. Finally, we have identified a new AMH target gene, Id3, which could be involved in the effects of AMH on the differentiation of granulosa cells and its other target cells.

  20. Optimization of culture media for large-scale lutein production by heterotrophic Chlorella vulgaris.

    Science.gov (United States)

    Jeon, Jin Young; Kwon, Ji-Sue; Kang, Soon Tae; Kim, Bo-Ra; Jung, Yuchul; Han, Jae Gap; Park, Joon Hyun; Hwang, Jae Kwan

    2014-01-01

    Lutein is a carotenoid with a purported role in protecting eyes from oxidative stress, particularly the high-energy photons of blue light. Statistical optimization was performed to growth media that supports a higher production of lutein by heterotrophically cultivated Chlorella vulgaris. The effect of media composition of C. vulgaris on lutein was examined using fractional factorial design (FFD) and central composite design (CCD). The results indicated that the presence of magnesium sulfate, EDTA-2Na, and trace metal solution significantly affected lutein production. The optimum concentrations for lutein production were found to be 0.34 g/L, 0.06 g/L, and 0.4 mL/L for MgSO4 ·7H2 O, EDTA-2Na, and trace metal solution, respectively. These values were validated using a 5-L jar fermenter. Lutein concentration was increased by almost 80% (139.64 ± 12.88 mg/L to 252.75 ± 12.92 mg/L) after 4 days. Moreover, the lutein concentration was not reduced as the cultivation was scaled up to 25,000 L (260.55 ± 3.23 mg/L) and 240,000 L (263.13 ± 2.72 mg/L). These observations suggest C. vulgaris as a potential lutein source.

  1. Antioxidant properties of lutein contribute to the protection against lipopolysaccharide-induced uveitis in mice

    Directory of Open Access Journals (Sweden)

    Yao Xin-Sheng

    2011-10-01

    Full Text Available Abstract Background Lutein is an important eye-protective nutrient. This study investigates the protective effects and mechanisms of lutein on lipopolysaccharides (LPS-induced uveitis in mice. Methods Lutein, suspended in drinking water at a final concentration of 12.5 and 25 mg/mL, was administered to mice at 0.1 mL/10 g body weight for five consecutive days. Control and model group received drinking water only. Uveitis was induced by injecting LPS (100 mg per mouse into the footpad in the model and lutein groups on day 5 after the last drug administration. Eyes of the mice were collected 24 hours after the LPS injection for the detection of indicators using commercial kits and reverse transcription-polymerase chain reaction. Results LPS-induced uveitis was confirmed by significant pathological damage and increased the nitric oxide level in eye tissue of BALB/C mice 24 hours after the footpad injection. The elevated nitric oxide level was significantly reduced by oral administration of lutein (125 and 500 mg/kg/d for five days before LPS injection. Moreover, lutein decreased the malondialdehyde content, increased the oxygen radical absorbance capacity level, glutathione, the vitamin C contents and total superoxide dismutase (SOD and glutathione peroxidase (GPx activities. Lutein further increased expressions of copper-zinc SOD, manganese SOD and GPx mRNA. Conclusion The antioxidant properties of lutein contribute to the protection against LPS-induced uveitis, partially through the intervention of inflammation process.

  2. A possible role for lutein and zeaxanthin in cognitive function in the elderly

    Science.gov (United States)

    Epidemiological studies suggest that dietary lutein and zeaxanthin may be of benefit in maintaining cognitive health. Among the carotenoids, lutein and zeaxanthin, are the only two that cross the blood-retina barrier to form macular pigment (MP) in the eye. They also preferentially accumulate in hum...

  3. Production, extraction and stabilization of lutein from microalga Chlorella sorokiniana MB-1.

    Science.gov (United States)

    Chen, Chun-Yen; Jesisca; Hsieh, Chienyan; Lee, Duu-Jong; Chang, Chien-Hsiang; Chang, Jo-Shu

    2016-01-01

    The efficiencies of extraction and preservation of lutein from microalgae are critical for the success of its commercialization. In this study, lutein was produced by Chlorella sorokiniana MB-1 via semi-batch mixotrophic cultivation. The microalgal biomass with a lutein content of 5.21mg/g was pretreated by bead-beating and high pressure cell disruption methods, and the lutein content was harvested by a reduced pressure extraction method. The effect of pretreatment, pressure, solvent type, extraction time and temperature on lutein recovery was investigated. Using high pressure pretreatment followed by extraction with tetrahydrofuran (THF) as solvent resulted in high lutein recovery efficiencies of 87.0% (20min) and 99.5% (40min) at 850mbar and 25°C. In contrast, using ethanol as the solvent, 86.2% lutein recovery was achieved under 450mbar, 35°C and 40min extraction. The extracted lutein was stabilized in olive oil or sunflower oil with half-lives of 53.1 and 63.8days, respectively. Copyright © 2015 Elsevier Ltd. All rights reserved.

  4. Ultrasound assisted extraction in quantifying lutein from chicken liver using high-performance liquid chromatography.

    Science.gov (United States)

    Sun, Ting; Xu, Zhimin; Godber, J Samuel

    2006-01-01

    Four sample preparation methods, (1) solvent (SOL), (2) saponification and solvent (SP), (3) ultrasound assisted solvent (UA), and (4) saponification and ultrasound assisted solvent (SP-UA), were used for quantifying lutein in chicken liver samples by HPLC. The lutein concentrations obtained by using SOL, UA, SP, and SP-UA were significantly different with values from 10.4 microg/g (UA) to undetected (SOL). Efficiency of the four different methods for extracting lutein from high to low were the UA, SP, SP-UA, and SOL method. The measured value of lutein in the liver sample using the UA method was approximately two and three times higher than that obtained from the SP and SP-UA method, respectively. The methods with saponification significantly affected the stabilities of lutein in liver samples. The lutein concentration measured with the solvent only method was either much lower than any of the other extraction methods or undetectable. This indicated that little lutein in those samples was in a form that could be extracted directly by solvent. Compared with the saponification method, the ultrasound assisted solvent method could effectively extract lutein from sample matrix and thus avoid chemical degradation reactions, which would be especially important for complex biological tissue such as liver.

  5. Genetic models for the study of luteinizing hormone receptor function

    Directory of Open Access Journals (Sweden)

    Prema eNarayan

    2015-09-01

    Full Text Available The luteinizing hormone/chorionic gonadotropin receptor, LHCGR, is essential for fertility in men and women. LHCGR binds luteinizing hormone (LH as well as the highly homologous chorionic gonadotropin (CG. Signaling from LHCGR is required for steroidogenesis and gametogenesis in males and females and for sexual differentiation in the male. The importance of LHCGR in reproductive physiology is underscored by the large number of naturally occurring inactivating and activating mutations in the receptor that result in reproductive disorders. Consequently, several genetically modified mouse models have been developed for the study of LHCGR function. They include targeted deletion of LH and LHCGR that mimic inactivating mutations in hormone and receptor, expression of a constitutively active mutant in LHCGR that mimics activating mutations associated with familial male-limited precocious puberty and transgenic models of LH and hCG overexpression. This review summarizes the salient findings from these models and their utility in understanding the physiological and pathological consequences of loss and gain of function in LHCGR signaling.

  6. Response of luteinizing hormone and follicle stimulating hormone to luteinizing hormone-releasing hormone in prepubertal and pubertal chidren, as measured by a highly sensitive immunradiometric assay

    OpenAIRE

    樋口,譲二

    1992-01-01

    To investigate the age-related changes in the pituitary responsiveness to luteinizing hormone-releasing hormone (LH-RH), the consentrations of serum luteinizing hormone (LH) and follicle stimulating hormone (FSH) were measured before and after LH-RH administra-tion using the highly sensitive immunoradiometric assay (IRMA) in 283 normal children (161 males and 77 females) between 4 and 14 years old and in 22 patients (18 males and 4 females) with pituitary dwarfism. Then, the area of response ...

  7. Astragalin, a Flavonoid from Morus alba (Mulberry Increases Endogenous Estrogen and Progesterone by Inhibiting Ovarian Granulosa Cell Apoptosis in an Aged Rat Model of Menopause

    Directory of Open Access Journals (Sweden)

    Min Wei

    2016-05-01

    Full Text Available Background: To determine the mechanism by which the flavonoid glycoside astragalin (AST reduces ovarian failure in an aged rat model of menopause. Methods: The in vivo effect of AST on granulosa cell (GC apoptosis in aged female rats was determined using flow cytometry. In vitro, the effects of AST on cultured GCs were investigated using the MTT proliferation assay and western blot assays. Results: Aged rats had significantly higher GC apoptosis as compared with young female rats. Treatment of aged rats with AST (all three doses; p < 0.01 or Progynova (p < 0.01 significantly reduced GC apoptosis as compared with the aged controls. The proportions of total apoptotic GCs was 25.70%, 86.65%, 47.04%, 27.02%, 42.09% and 56.42% in the normal, aged, 17β-estradiol (E2, high dose AST, medium dose AST, and low dose AST-treated groups, respectively. Significant increases of serum E2 and P4 levels, as well as altered levels of serum follicle stimulating hormone (FSH and luteinizing hormone (LH levels. In cultured rat GCs, AST stimulated GC proliferation, E2 and progesterone (P4 secretion, reduced apoptosis, reduced the level of the pro-apoptotic protein Bcl-2 (p < 0.01, but had no effect on BAX. Conclusions: AST enhanced ovarian function in aged female rats by increasing E2 and P4 levels, and reducing ovarian GC apoptosis via a mechanism involving Bcl-2. These data demonstrate a new pharmacological activity for AST, as well as a novel mechanism of action, and further suggest that AST may be a new therapeutic agent for the management of menopausal symptoms.

  8. An incidental ovarian mass: A case of ovarian hemangioma with prominent stromal luteinization

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    Babak Shirazi

    2015-01-01

    Full Text Available Ovarian hemangioma is a rare benign tumor of female genital tract. Stromal luteinization in ovarian hemangioma is an uncommon process and the pathogenesis is controversial. In this regard, two hypotheses have been suggested whether luteinization is a reactive process or it is the stimulator for development of ovarian hemangioma. Here, we report a case of a 55-year-old woman who referred to our center due to incidental finding of left ovarian mass in pelvic sonography. Microscopically, the mass showed a mixed cavernous and capillary hemangioma and the peripheral stroma contained several small and large clusters of stromal cells, which were luteinized. It should be noted that an ovarian hemangioma could be associated with stromal luteinization although its pathogenesis is not clearly known. Yet, we believe the stromal luteinization around ovarian hemangioma could be a reactive phenomenon.

  9. The LIM domain protein FHL2 interacts with the NR5A family of nuclear receptors and CREB to activate the inhibin-α subunit gene in ovarian granulosa cells.

    Science.gov (United States)

    Matulis, Christina K; Mayo, Kelly E

    2012-08-01

    Nuclear receptor transcriptional activity is enhanced by interaction with coactivators. The highly related nuclear receptor 5A (NR5A) subfamily members liver receptor homolog 1 and steroidogenic factor 1 bind to and activate several of the same genes, many of which are important for reproductive function. To better understand transcriptional activation by these nuclear receptors, we sought to identify interacting proteins that might function as coactivators. The LIM domain protein four and a half LIM domain 2 (FHL2) was identified as interacting with the NR5A receptors in a yeast two-hybrid screen of a human ovary cDNA library. FHL2, and the closely related FHL1, are both expressed in the rodent ovary and in granulosa cells. Small interfering RNA-mediated knockdown of FHL1 and FHL2 in primary mouse granulosa cells reduced expression of the NR5A target genes encoding inhibin-α and P450scc. In vitro assays confirmed the interaction between the FHL and NR5A proteins and revealed that a single LIM domain of FHL2 is sufficient for this interaction, whereas determinants in both the ligand binding domain and DNA binding domain of NR5A proteins are important. FHL2 enhances the ability of both liver receptor homolog 1 and steroidogenic factor 1 to activate the inhibin-α subunit gene promoter in granulosa cells and thus functions as a transcriptional coactivator. FHL2 also interacts with cAMP response element-binding protein and substantially augments activation of inhibin gene expression by the combination of NR5A receptors and forskolin, suggesting that FHL2 may facilitate integration of these two signals. Collectively these results identify FHL2 as a novel coactivator of NR5A nuclear receptors in ovarian granulosa cells and suggest its involvement in regulating target genes important for mammalian reproduction.

  10. effect of luteinizing hormone-releasing hormone analogue on the sexual behavior of sacalia quadriocellata

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    luteinizing hormone-releasing hormone (lhrh) is known to influence sexual behavior in many vertebrate taxa,but there have been no systematic studies on the role of lhrh in sexual behavior of turtles.we tested the hypotheses that exogenous lhrh analogues would induce sexual behavior of male four-eyed turtle,sacalia quadriocellata.we examined this by challenging males with intramuscular injections of mammalian luteinizing hormone-releasing hormone analogue (lhrh-a),human chorionic gonadotropin (hcg),or a combination of the two,and subsequently exposing them to sexually receptive females for behavioral observation.our data show that the injection of only hcg could not,while that of only lhrh-a could,facilitate sexual behavior along with testicular recrudescence and spermatogenesis in s.quadriocellata.the injection of both lhrh-a and hcg would induce more drastic sexual behavior of the animals than that of lhrh-a alone,indicating hcg enhances the effects of lhrh-a induced sexual behavior.however,different pharmacological dosages of lhrh-a (0.5 μg,1 μg,2 μg per 100 g bodyweight) did not correspond to different activity levels.though the mechanism of lhrh effect was not determined,this study may support that the sexual behavior ofs.quadriocellata which occurs at the beginning of the injection despite regression of the gonads.this is the first report on the exogenous lhrh-a induced sexual behavior for this species.

  11. Physicochemical properties and storage stability of lutein microcapsules prepared with maltodextrins and sucrose by spray drying.

    Science.gov (United States)

    Kuang, Pengqun; Zhang, Hongchao; Bajaj, Poonam R; Yuan, Qipeng; Tang, Juming; Chen, Shulin; Sablani, Shyam S

    2015-02-01

    The purpose of this study was to determine the physicochemical properties of lutein microcapsules. Nine types of lutein microcapsules were prepared in order to determine their encapsulation efficiency and yield. Results show that lutein microcapsules with maltodextrin M040 and sucrose at the weight ratio of 3:1 (designated as M040:1) had the highest encapsulation efficiency (90.1%) among the lutein microcapsules, as well as a higher encapsulation yield (90.4%). The onset glass transition temperatures (Tgi ) and the surface dents of the lutein microcapsules decreased as the dextrose equivalent value of maltodextrin and the weight ratio of sucrose increased. Enthalpy relaxation experiments were conducted for the lutein microcapsules M040:1 at (Tgi - 5) , (Tgi - 10), and (Tgi - 15) °C, and the obtained data were fitted to the Kohlrausch-Williams-Watts model. Results show that the mean relaxation time (τ) (316 h) of M040:1 lutein microcapsules aged at (Tgi - 15) °C was greater than the τ (161 h) at (Tgi - 10) °C and τ (60.5 h) at (Tgi - 5) °C. Effects of temperature and oxygen transmission rates for package film on the storage stability of M040:1 lutein microcapsules were also investigated. Findings show that rates of lutein degradation and color change increased by an order of magnitude as storage temperature (4 to 97 °C) and oxygen transmission rate of the package film (0.018 to 62.8 cc/m(2) day) increased. These results suggest that lutein is highly unstable and susceptible to thermal and oxidative degradations. However, microencapsulation with appropriate wall materials of higher relaxation time and high oxygen barrier packaging can increase the storage life.

  12. Further studies of the effects of follicular fluid and membrana granulosa cells on the spontaneous maturation of pig oocytes.

    Science.gov (United States)

    Racowsky, C; McGaughey, R W

    1982-11-01

    Liberated cumulus-enclosed pig oocytes were cultured either alone in follicular fluid or with membrana granulosa cells in a complex serum based medium. After 24 h, oocytes were air-dried for cytogenetic analysis, meiotic stage was scored, and viability of granulosa cells was determined. Neither the release from meiotic arrest nor the progression of maturation to metaphase II was significantly inhibited by either of these follicular components. Co-culture of membrana granulosa cells and oocytes significantly stimulated maturation in one experimental series, while viability of the somatic cells was maintained in all experiments. These results do not support the concept of a stable oocyte maturation inhibitor of granulosa cell origin in follicular fluid.

  13. Cellular heterogeneity in the membrana granulosa of developing rat follicles: assessment by flow cytometry and lectin binding.

    Science.gov (United States)

    Kerketze, K; Blaschuk, O W; Farookhi, R

    1996-07-01

    The hormone-mediated maturation of ovarian follicles is apparently accompanied by position-specific differentiation of cells of the membrana granulosa. We have assessed the extent of this cellular heterogeneity by flow cytometry using a variety of fluorescein isothiocyanate-labeled lectins as probes. Follicular development was stimulated in immature rats by treatment with either diethylstilbestrol (DES) or equine CG (eCG). Lectin binding to monodispersed rat granulosa cells was then analyzed by flow cytometry. Our results demonstrate that there are two distinct populations of small (4-7 microM) and large (9-12 microM) granulosa cells in follicles from DES- and eCG-treated animals. Both populations appear to be mitotically active and show specific lectin-binding characteristics. Six lectins (canavalia ensiforms, triticum vulgaris, maclura pomifera, erythrina cristagalli, jacalin, and vicia villosa) bind equally to both small and large granulosa cells from the DES- and eCG-treated rats. In contrast, no binding to either cell population was detected with six other lectins (dolichos biflorus, griffonia simplicifolia-II, lycopersicon esculentum, datura stramonium, solanum tuberosum, and ulex europaeus). Furthermore, four galactose-binding lectins (bauhinia purpurea, glysine maximus, griffonia simplicifolia-I, and arachis hypogaea) were found to identify specific subsets of granulosa cells. Three of these lectins (bauhinia purpurea, glysine maximus, and griffonia simplicifolia-I) bind to only small granulosa cells from either DES- or eCG- treated immature rats. The fourth lectin (arachis hypogaea) identifies subpopulations of both small and large granulosa cells. Application of the four galactose-specific lectins to fixed sections of frozen ovaries demonstrated binding to the perioocyte and cumulus granulosa cells. We conclude that cellular heterogeneity exists within the follicular epithelium at various stages-specific lectin-binding sites.

  14. Comparison of the effects of human and chicken ghrelin on chicken ovarian hormone release.

    Science.gov (United States)

    Sirotkin, Alexander V; Harrath, Abdel Halim; Grossmann, Roland

    2016-11-01

    The aim of the present experiments was to examine the species-specific and cell-specific effects of ghrelin on chicken ovarian hormone release. For this purpose, we compared the effects of chicken and human ghrelin on the release of estradiol (E), testosterone (T), progesterone (P) and arginine-vasotocin (AVT) by cultured fragments of chicken ovarian follicles and on the release of T and AVT by cultured ovarian granulosa cells. In cultured chicken ovarian fragments, both human and chicken ghrelin promoted E release. T output was stimulated by chicken ghrelin but not by human ghrelin. No effect of either human or chicken ghrelin on P release was observed. Human ghrelin promoted but chicken ghrelin suppressed AVT release by chicken ovarian fragments. In cultured ovarian granulosa cells, human ghrelin inhibited while chicken ghrelin stimulated T release. Both human and chicken ghrelin suppressed AVT output by chicken granulosa cells. These data confirm the involvement of ghrelin in the control of ovarian secretory activity and demonstrate that the effect of ghrelin is species-specific. The similarity of avian ghrelin on avian ovarian granulosa cells and ovarian fragments (containing both granulosa and theca cells) suggests that ghrelin can influence chicken ovarian hormones primarily by acting on granulosa cells.

  15. Prostaglandin E2 (EP) receptors mediate PGE2-specific events in ovulation and luteinization within primate ovarian follicles.

    Science.gov (United States)

    Kim, Soon Ok; Harris, Siabhon M; Duffy, Diane M

    2014-04-01

    Prostaglandin E2 (PGE2) is a key mediator of ovulation. All 4 PGE2 receptors (EP receptors) are expressed in the primate follicle, but the specific role of each EP receptor in ovulatory events is poorly understood. To examine the ovulatory events mediated via these EP receptors, preovulatory monkey follicles were injected with vehicle, the PG synthesis inhibitor indomethacin, or indomethacin plus PGE2. An ovulatory dose of human chorionic gonadotropin was administered; the injected ovary was collected 48 hours later and serially sectioned. Vehicle-injected follicles showed normal ovulatory events, including follicle rupture, absence of an oocyte, and thickening of the granulosa cell layer. Indomethacin-injected follicles did not rupture and contained oocytes surrounded by unexpanded cumulus; granulosa cell hypertrophy did not occur. Follicles injected with indomethacin plus PGE2 were similar to vehicle-injected ovaries, indicating that PGE2 restored the ovulatory changes inhibited by indomethacin. Additional follicles were injected with indomethacin plus an agonist for each EP receptor. EP1, EP2, and EP4 agonists each promoted aspects of follicle rupture, but no single EP agonist recapitulated normal follicle rupture as seen in follicles injected with either vehicle or indomethacin plus PGE2. Although EP4 agonist-injected follicles contained oocytes in unexpanded cumulus, the absence of oocytes in EP1 agonist- and EP2 agonist-injected follicles suggests that these EP receptors promote cumulus expansion. Surprisingly, the EP3 agonist did not stimulate any of these ovulatory changes, despite the high level of EP3 receptor expression in the monkey follicle. Therefore, agonists and antagonists selective for EP1 and EP2 receptors hold the most promise for control of ovulatory events in women.

  16. Critical Role of FoxO1 in Granulosa Cell Apoptosis Caused by Oxidative Stress and Protective Effects of Grape Seed Procyanidin B2

    OpenAIRE

    Jia-Qing Zhang; Bin-Wen Gao; Jing Wang; Qiao-Ling Ren; Jun-Feng Chen; Qiang Ma; Zi-Jing Zhang; Bao-Song Xing

    2016-01-01

    Reactive oxygen species (ROS) are closely related to the follicular granulosa cell apoptosis. Grape seed procyanidin B2 (GSPB2) has been reported to possess potent antioxidant activity. However, the GSPB2-mediated protective effects and the underlying molecular mechanisms in granulosa cell apoptosis process remain unknown. In this study, we showed for the first time that GSPB2 treatment decreased FoxO1 protein level, improved granulosa cell viability, upregulated LC3-II protein level, and red...

  17. Luteinizing hormone promotes gonadal tumorigenesis in inhibin-deficient mice

    Science.gov (United States)

    Nagaraja, Ankur K.; Agno, Julio E.; Kumar, T. Rajendra; Matzuk, Martin M.

    2009-01-01

    Summary The inhibins are secreted α:β heterodimers of the TGF-β superfamily that are mainly synthesized in Sertoli cells and granulosa cells, and are critical regulators of testicular and ovarian development and function. Mice homozygous for a targeted deletion of the inhibin α subunit gene (Inha-/-) develop sex cord-stromal tumors in a gonadotropin-dependent manner. Here, we determine the contribution of LH to gonadal tumorigenesis by generating mice deficient in both inhibins and LH. Inha-/-Lhb-/- mice have increased survival and delayed tumor progression, and these observations correlate with lower serum FSH and estradiol levels compared to Inha-/- controls. Double mutant testicular tumors demonstrate decreased expression of cyclin D2, while double mutant ovarian tumors have elevated expression of p15INK4b and trend toward higher levels of p27Kip1. We conclude that LH is not required for tumor formation in the absence of inhibins but promotes tumor progression, likely through alterations in serum hormone levels and cell cycle regulators. PMID:18657590

  18. Effects of Lutein on the Growth and Migration of Bovine Lens Epithelial Cells In Vitro

    Institute of Scientific and Technical Information of China (English)

    Yizhen HU; Zhirong XU

    2008-01-01

    The effects of lutein on the growth and migration of bovine lens epithelial cells (BLECs) in vitro were observed in an attempt to find a drug that can prevent after-cataract. BLECs were cultured in vitro and different concentrations of lutein were added to the BLECs cultures of the second and third generations. The effects of lutein on the proliferation of BLECs in vitro were examined by the MTT method, and the migration of BLECs was evaluated by a scratch wound assay. The results showed that: (1) Lutein at concentrations of 1 to 16μmol/L could inhibit the proliferation of BLECs in a dose-and time-dependent manner (P<0.01); (2) The migration of BLECs was evaluated by wound healing rate. As compared with the control group, the wound healing rate in the experimental groups was decreased from 0.672±0.164 to -0.234±0.144 and -0.597±0.063 (P<0.01) at 1 and 2μmol/L lutein, respectively. It was concluded that lutein at concentration of ≥1μmol/L inhibited the proliferation and migration of BLECs in vitro. Lutein may become an effective drug to prevent after-cataract.

  19. EFSA Panel on Dietetic Products, Nutrition and Allergies (NDA); Scientific Opinion on the substantiation of health claims related to lutein and maintenance of normal vision (ID 1603, 1604, further assessment) pursuant to Article 13(1) of Regulation (EC) No 1924/2006

    DEFF Research Database (Denmark)

    Tetens, Inge

    Following a request from the European Commission, the Panel on Dietetic Products, Nutrition and Allergies was asked to provide a scientific opinion on a list of health claims pursuant to Article 13 of Regulation (EC) No 1924/2006 in the framework of further assessment related to lutein and mainte......Following a request from the European Commission, the Panel on Dietetic Products, Nutrition and Allergies was asked to provide a scientific opinion on a list of health claims pursuant to Article 13 of Regulation (EC) No 1924/2006 in the framework of further assessment related to lutein...... into account that one human intervention study in healthy subjects did not show an effect of lutein on visual acuity or glare sensitivity, that the results of this study were inconsistent as regards contrast sensitivity, and that the evidence provided for a mechanism by which lutein could exert the claimed...

  20. Elevation of lutein content in tomato: a biochemical tug-of-war between lycopene cyclases.

    Science.gov (United States)

    Giorio, Giovanni; Yildirim, Arzu; Stigliani, Adriana Lucia; D'Ambrosio, Caterina

    2013-11-01

    Lutein is becoming increasingly important in preventive medicine due to its possible role in maintaining good vision and in preventing age-related maculopathy. Average daily lutein intake in developed countries is often below suggested daily consumption levels, and lutein supplementation could be beneficial. Lutein is also valuable in the food and feed industries and is emerging in nutraceutical and pharmaceutical markets. Currently, lutein is obtained at high cost from marigold petals, and synthesis alternatives are thus desirable. Tomato constitutes a promising starting system for production as it naturally accumulates high levels of lycopene. To develop tomato for lutein synthesis, the tomato Red Setter cultivar was transformed with the tomato lycopene ε-cyclase-encoding gene under the control of a constitutive promoter, and the HighDelta (HD) line, characterised by elevated lutein and δ-carotene content in ripe fruits, was selected. HD was crossed to the transgenic HC line and to RS(B) with the aim of converting all residual fruit δ-carotene to lutein. Fruits of both crosses were enriched in lutein and presented unusual carotenoid profiles. The unique genetic background of the crosses used in this study permitted an unprecedented analysis of the role and regulation of the lycopene cyclase enzymes in tomato. A new defined biochemical index, the relative cyclase activity ratio, was used to discern post-transcriptional regulation of cyclases, and will help in the study of carotenoid biosynthesis in photosynthetic plant species and particularly in those, like tomato, that have been domesticated for the production of food, feed or useful by-products.

  1. Enhancement of Lutein Production in Chlorella sorokiniana (Chorophyta by Improvement of Culture Conditions and Random Mutagenesis

    Directory of Open Access Journals (Sweden)

    Maria Angeles Vargas

    2011-09-01

    Full Text Available Chlorella sorokiniana has been selected for lutein production, after a screening of thirteen species of microalgae, since it showed both a high content in this carotenoid and a high growth rate. The effects of several nutritional and environmental factors on cell growth and lutein accumulation have been studied. Maximal specific growth rate and lutein content were attained at 690 µmol photons m−2 s−1, 28 °C, 2 mM NaCl, 40 mM nitrate and under mixotrophic conditions. In general, optimal conditions for the growth of this strain also lead to maximal lutein productivity. High lutein yielding mutants of C. sorokiniana have been obtained by random mutagenesis, using N-methyl-N′-nitro-nitrosoguanidine (MNNG as a mutagen and selecting mutants by their resistance to the inhibitors of the carotenogenic pathway nicotine and norflurazon. Among the mutants resistant to the herbicides, those exhibiting both high content in lutein and high growth rate were chosen. Several mutants exhibited higher contents in this carotenoid than the wild type, showing, in addition, either a similar or higher growth rate than the latter strain. The mutant MR-16 exhibited a 2.0-fold higher volumetric lutein content than that of the wild type, attaining values of 42.0 mg L−1 and mutants DMR-5 and DMR-8 attained a lutein cellular content of 7.0 mg g−1 dry weight. The high lutein yield exhibited by C. sorokiniana makes this microalga an excellent candidate for the production of this commercially interesting pigment.

  2. Dephosphorylation and inactivation of NPR2 guanylyl cyclase in granulosa cells contributes to the LH-induced decrease in cGMP that causes resumption of meiosis in rat oocytes

    Science.gov (United States)

    Egbert, Jeremy R.; Shuhaibar, Leia C.; Edmund, Aaron B.; Van Helden, Dusty A.; Robinson, Jerid W.; Uliasz, Tracy F.; Baena, Valentina; Geerts, Andreas; Wunder, Frank; Potter, Lincoln R.; Jaffe, Laurinda A.

    2014-01-01

    In mammals, the meiotic cell cycle of oocytes starts during embryogenesis and then pauses. Much later, in preparation for fertilization, oocytes within preovulatory follicles resume meiosis in response to luteinizing hormone (LH). Before LH stimulation, the arrest is maintained by diffusion of cyclic (c)GMP into the oocyte from the surrounding granulosa cells, where it is produced by the guanylyl cyclase natriuretic peptide receptor 2 (NPR2). LH rapidly reduces the production of cGMP, but how this occurs is unknown. Here, using rat follicles, we show that within 10 min, LH signaling causes dephosphorylation and inactivation of NPR2 through a process that requires the activity of phosphoprotein phosphatase (PPP)-family members. The rapid dephosphorylation of NPR2 is accompanied by a rapid phosphorylation of the cGMP phosphodiesterase PDE5, an enzyme whose activity is increased upon phosphorylation. Later, levels of the NPR2 agonist C-type natriuretic peptide decrease in the follicle, and these sequential events contribute to the decrease in cGMP that causes meiosis to resume in the oocyte. PMID:25183874

  3. Dephosphorylation and inactivation of NPR2 guanylyl cyclase in granulosa cells contributes to the LH-induced decrease in cGMP that causes resumption of meiosis in rat oocytes.

    Science.gov (United States)

    Egbert, Jeremy R; Shuhaibar, Leia C; Edmund, Aaron B; Van Helden, Dusty A; Robinson, Jerid W; Uliasz, Tracy F; Baena, Valentina; Geerts, Andreas; Wunder, Frank; Potter, Lincoln R; Jaffe, Laurinda A

    2014-09-01

    In mammals, the meiotic cell cycle of oocytes starts during embryogenesis and then pauses. Much later, in preparation for fertilization, oocytes within preovulatory follicles resume meiosis in response to luteinizing hormone (LH). Before LH stimulation, the arrest is maintained by diffusion of cyclic (c)GMP into the oocyte from the surrounding granulosa cells, where it is produced by the guanylyl cyclase natriuretic peptide receptor 2 (NPR2). LH rapidly reduces the production of cGMP, but how this occurs is unknown. Here, using rat follicles, we show that within 10 min, LH signaling causes dephosphorylation and inactivation of NPR2 through a process that requires the activity of phosphoprotein phosphatase (PPP)-family members. The rapid dephosphorylation of NPR2 is accompanied by a rapid phosphorylation of the cGMP phosphodiesterase PDE5, an enzyme whose activity is increased upon phosphorylation. Later, levels of the NPR2 agonist C-type natriuretic peptide decrease in the follicle, and these sequential events contribute to the decrease in cGMP that causes meiosis to resume in the oocyte.

  4. In vitro and in vivo effects of lutein against cisplatin-induced ototoxicity.

    Science.gov (United States)

    Roldán-Fidalgo, A; Martín Saldaña, S; Trinidad, A; Olmedilla-Alonso, B; Rodríguez-Valiente, A; García-Berrocal, J R; Ramírez-Camacho, R

    2016-04-01

    Cisplatin is a commonly prescribed drug that produces ototoxicity as a side effect. Lutein is a carotenoid with antioxidant and anti-inflammatory properties previously tested for eye, heart and skin diseases but not evaluated to date in ear diseases. To evaluate the protective effects of lutein on HEI-OC1 auditory cell line and in a Wistar rat model of cisplatin ototoxicity. In vitro study: Culture HEI-OC1 cells were exposed to lutein (2.5-100 μM) and to 25 μM cisplatin for 24h. In vivo study: Twenty eight female Wistar rats were randomized into three groups. Group A (n=8) received intratympanic lutein (0.03 mL) (1mg/mL) in the right ear and saline solution in the left one to determine the toxicity of lutein. Group B (n=8) received also intraperitoneal cisplatin (10mg/kg) to test the efficacy of lutein against cisplatin ototoxicity. Group C (n=12) received intratympanic lutein (0.03 mL) (1mg/mL) to quantify lutein in cochlear fluids (30 min, 1h and 5 days after treatment). Hearing function was evaluated by means of Auditory Steady-State Responses before the procedure and 5 days after (groups A and B). Morphological changes were studied by confocal laser scanning microscopy. In vitro study: Lutein significantly reduced the cisplatin-induced cytotoxicity in the HEI-OC1 cells when they were pre-treated with lutein concentrations of 60 and 80 μM. In vivo study: Intratympanic lutein (1mg/mL) application showed no ototoxic effects. However it did not achieve protective effect against cisplatin-induced ototoxicity in Wistar rats. Although lutein has shown beneficial effects in other pathologies, the present study only obtained protection against cisplatin ototoxicity in culture cells, but not in the in vivo model. The large molecule size, the low dose administered, and restriction to diffusion in the inner ear could account for this negative result. Copyright © 2016 Elsevier GmbH. All rights reserved.

  5. Antioxidant properties of lutein contribute to the protection against lipopolysaccharide-induced uveitis in mice

    OpenAIRE

    Yao Xin-Sheng; Yao Nan; Lan Fang; Tsoi Bun; He Rong-Rong; Kurihara Hiroshi

    2011-01-01

    Abstract Background Lutein is an important eye-protective nutrient. This study investigates the protective effects and mechanisms of lutein on lipopolysaccharides (LPS)-induced uveitis in mice. Methods Lutein, suspended in drinking water at a final concentration of 12.5 and 25 mg/mL, was administered to mice at 0.1 mL/10 g body weight for five consecutive days. Control and model group received drinking water only. Uveitis was induced by injecting LPS (100 mg per mouse) into the footpad in the...

  6. Antioxidant properties of lutein contribute to the protection against lipopolysaccharide-induced uveitis in mice

    OpenAIRE

    He, Rong-rong; Tsoi, Bun; Lan, Fang; Yao, Nan; Yao, Xin-Sheng; Kurihara, Hiroshi

    2011-01-01

    Background Lutein is an important eye-protective nutrient. This study investigates the protective effects and mechanisms of lutein on lipopolysaccharides (LPS)-induced uveitis in mice. Methods Lutein, suspended in drinking water at a final concentration of 12.5 and 25 mg/mL, was administered to mice at 0.1 mL/10 g body weight for five consecutive days. Control and model group received drinking water only. Uveitis was induced by injecting LPS (100 mg per mouse) into the footpad in the model an...

  7. The Transcriptional Targets of Mutant FOXL2 in Granulosa Cell Tumours

    OpenAIRE

    2012-01-01

    BACKGROUND: Despite their distinct biology, granulosa cell tumours (GCTs) are treated the same as other ovarian tumours. Intriguingly, a recurring somatic mutation in the transcription factor Forkhead Box L2 (FOXL2) 402C>G has been found in nearly all GCTs examined. This investigation aims to identify the pathogenicity of mutant FOXL2 by studying its altered transcriptional targets. METHODS: The expression of mutant FOXL2 was reduced in the GCT cell line KGN, and wildtype and mutant FOXL2 wer...

  8. Pig membrana granulosa cells prevent resumption of meiosis in cattle oocytes.

    Science.gov (United States)

    Kalous, J; Sutovsky, P; Rimkevicova, Z; Shioya, Y; Lie, B L; Motlik, J

    1993-01-01

    Membrana granulosa was isolated from healthy large antral follicles of prepubertal or cyclic gilts stimulated with PMSG or PMSG and hCG. Ultrastructural observations revealed that pieces of pig membrana granulosa were associated with the basement membrane. The cattle cumulus-enclosed oocytes (COC) were placed in the rolled pieces of the pig membrana granulosa (PMG). After 8 and 24 hr of coculture with PMG from prepubertal gilts, only 16% and 21% of oocytes underwent GVBD, respectively. PMG from PMSG-stimulated cyclic gilts blocked the resumption of meiosis in all COC. The inhibitory effect of heterologous granulosa cells was fully reversible. When COC were initially incubated for 2 and 4 hr, subsequent culture in PMG prevented GVBD in 100% and 36% of oocytes, respectively. This suggests that functional contact between COC and PMG was established during the first 2 hr of coculture. To follow metabolic cooperation between PMG and COC, PMG was prelabeled with 3H-uridine and cocultured with COC. Autoradiography on semithin sections revealed the intensive passage of 3H-uridine from PMG into the cumulus layer and an oocyte. COC placed in PMG after GVBD (8 and 12 hr of an initial incubation) did not extrude the first polar body. PMG isolated from cyclic gilts after PMSG and hCG stimulation also inhibited GVBD of COC. Since nearly all COC placed in PMG isolated 10 and 12 hr after hCG remained in the GV stage after 24 hr of coculture, the hCG stimulation did not substantially diminish the meiosis inhibiting activity of PMG.(ABSTRACT TRUNCATED AT 250 WORDS)

  9. NO-mediated regulation of GLUT by T3 and FSH in rat granulosa cells.

    Science.gov (United States)

    Tian, Ye; Ding, Yu; Liu, Juan; Heng, Dai; Xu, Kaili; Liu, Wenbo; Zhang, Cheng

    2017-03-17

    Thyroid hormones (THs) are important for normal reproductive function. Although 3,5,3'-triiodothyronine (T3) enhances follicle-stimulating hormone (FSH)-induced preantral follicle growth and granulosa cells development in vitro, little is known about the molecular mechanisms regulating ovarian development via glucose. In this study, we investigated whether and how T3 combines with FSH to regulate glucose transporter protein (GLUT) expression and glucose uptake in granulosa cells. Here, we present evidence that T3 and FSH co-treatment significantly increased GLUT-1/GLUT-4 expression, and translocation in cells, as well as glucose uptake. These changes were accompanied by upregulation of NOS3 expression, total NOS and NOS3 activity and NO content in granulosa cells. Furthermore, we found that activation of the mTOR and PI3K/Akt pathway is required for the regulation of GLUT expression, translocation, and glucose uptake by hormones. We also found that L-arginine (L-arg) up-regulated GLUT-1/GLUT-4 expression and translocation, which were related to increased glucose uptake, however, these responses were significantly blocked by L-NAME. In addition, inhibiting NO production attenuated T3 and FSH-induced GLUT expression, translocation, and glucose uptake in granulosa cells. Our data demonstrate that T3 and FSH co-treatment potentiates cellular glucose uptake via GLUT upregulation and translocation, which are mediated through the activation of the mTOR/PI3K/Akt pathway. Meanwhile, NOS3/NO are also involved in this regulatory system. These findings suggest that GLUT is a novel mediator of T3 and FSH-induced follicular development.

  10. Abnormal Mitochondrial Function and Impaired Granulosa Cell Differentiation in Androgen Receptor Knockout Mice

    Directory of Open Access Journals (Sweden)

    Ruey-Sheng Wang

    2015-04-01

    Full Text Available In the ovary, the paracrine interactions between the oocyte and surrounded granulosa cells are critical for optimal oocyte quality and embryonic development. Mice lacking the androgen receptor (AR−/− were noted to have reduced fertility with abnormal ovarian function that might involve the promotion of preantral follicle growth and prevention of follicular atresia. However, the detailed mechanism of how AR in granulosa cells exerts its effects on oocyte quality is poorly understood. Comparing in vitro maturation rate of oocytes, we found oocytes collected from AR−/− mice have a significantly poor maturating rate with 60% reached metaphase II and 30% remained in germinal vesicle breakdown stage, whereas 95% of wild-type AR (AR+/+ oocytes had reached metaphase II. Interestingly, we found these AR−/− female mice also had an increased frequency of morphological alterations in the mitochondria of granulosa cells with reduced ATP generation (0.18 ± 0.02 vs. 0.29 ± 0.02 µM/mg protein; p < 0.05 and aberrant mitochondrial biogenesis. Mechanism dissection found loss of AR led to a significant decrease in the expression of peroxisome proliferator-activated receptor γ (PPARγ co-activator 1-β (PGC1-β and its sequential downstream genes, nuclear respiratory factor 1 (NRF1 and mitochondrial transcription factor A (TFAM, in controlling mitochondrial biogenesis. These results indicate that AR may contribute to maintain oocyte quality and fertility via controlling the signals of PGC1-β-mediated mitochondrial biogenesis in granulosa cells.

  11. Ruptured Granulosa Cell Tumor of the Ovary as a Cause of Acute Abdomen in Postmenopausal Woman

    Directory of Open Access Journals (Sweden)

    Tufan Oge

    2012-01-01

    Full Text Available Acute abdomen with hemoperitoneum is a very rare entity in postmenopausal women due to gynecologic conditions. A 54-year-old, postmenopausal woman was brought to emergency department with severe abdominal pain. Physical examination revealed acute abdomen findings with 15 cm pelvic mass on the right adnexal region. Immediate exploratory laparotomy was performed. During laparotomy 1000 cc of bloodstained fluid, ruptured and actively bleeding large mass arising from right ovary was observed. Right salpingo-oopherectomy was performed in emergency conditions, and pathology report revealed an adult type of granulosa cell tumor. After this result, staging surgery was performed and patient was diagnosed as granulosa cell tumor stage 1 c. Cisplatin, etoposide, and bleomycin chemotherapy was given. Clinicians should be aware of granulosa cell tumors which may occur at any age and prone to rupture. Frozen section will be helpful in order to avoid incomplete surgeries especially in postmenopausal women presented with intra-abdominal bleeding.

  12. Immunogold study on lectin binding in the porcine zona pellucida and granulosa cells

    Directory of Open Access Journals (Sweden)

    F Parillo

    2009-06-01

    Full Text Available An ultrastructural localization of lectin receptors on the zona pellucida (ZP of porcine antral oocytes and on the granulosa cells was performed using a panel of horseradish peroxidase- labelled lectins in conjunction with antiperoxidase antibody and protein A-gold. In some cases, lectin incubation was preceded by sialidase digestion. WGA-, Con-A-, UEA-I-, RCA-I-, PNA- and SBA-reactive sites were distributed differently in the porcine ZP. Sialidase digestion increased the positivity obtained with RCA-I and it was necessary to promote PNA and SBA reactivity. These results indicated that the ZP contained N-acetylglucosamine, a-mannose, a- fucose, b-Gal-(1-4GlcNAc, b-Gal- (1-3GalNAc, b-GalNAc and sialic acid residues. We also observed the presence of vesicles in both the ooplasm and granulosa cells, showing a similar lectin binding pattern to that of the ZP, thus suggesting that the oocyte and granulosa cells are the site of synthesis of ZP glucidic determinants.

  13. Analysis of gene expression in granulosa cells of ovine antral growing follicles using suppressive subtractive hybridization.

    Science.gov (United States)

    Chen, A Qin; Wang, Zheng Guang; Xu, Zi Rong; Yu, Song Dong; Yang, Zhi Gang

    2009-10-01

    Follicular growth, development and ovulation are highly ordered processes that involve the expression of many genes under precise temporal and spatial regulation. However, information on stage-specific gene expression during the antral follicle phase in sheep is not well understood. In the present study, suppressive subtractive hybridization (SSH) was performed to screen genes that were differentially expressed in the granulosa cells between large follicles (LF, >5mm) and small follicles (SF, 3-5mm), and subtractive cDNA library was constructed. Furthermore, with dot-blot analysis, a total of 90 clones randomly selected from the library were proven to be differentially expressed in the granulosa cells. Among these, 38 exhibited high homology to known genes, 14 sequences were corresponding to novel expressed sequence tags (ESTs). Four ESTs, LAPTM4A, SERPINE2, GSTA1, and INHBA, were further examined the reproducibility of the SSH data by the real-time quantitative PCR. Results confirmed an increase expression of respective mRNA in granulosa cells of large follicles compared with that of small follicles. It is concluded that we have identified several genes (known or unknown) that may effect follicular growth, dominance or ovulation in ewes.

  14. Goose broodiness is involved in granulosa cell autophagy and homeostatic imbalance of follicular hormones.

    Science.gov (United States)

    Yu, Jing; Lou, Yaping; He, Ke; Yang, Songbai; Yu, Wensai; Han, Lu; Zhao, Ayong

    2016-05-01

    Broodiness is observed in most domestic fowls and influences egg production. The goose is one of the most important waterfowls, having strong broody behavior. However, whether autophagy and follicular internal environment play a role in the broodiness behavior of goose is unknown. In this report, we analyzed the follicular internal environment and granulosa cell autophagy of goose follicles. The results show that the contents of hormones, including prolactin (PRL), progesterone (P4), and estradiol (E2), increased in broody goose follicles. Most importantly, the level of granulosa cell autophagy in broody goose follicles was elevated, detected by electron microscopy and western blotting. Also, the expressions of positive regulators of autophagy, including miR-7, miR-29, miR-100, miR-181, PRLR, LC3, p53,Beclin1, Atg9, and Atg12, were up-regulated and the expressions of negative regulators of autophagy, including miR-34b and miR-34c, were down-regulated in broody goose follicles. Our results suggest that goose broodiness is involved in increased granulosa cell autophagy and homeostasis imbalance of internal environment in the follicles. This work contributes to our knowledge of goose broodiness and may influence egg production. © 2016 Poultry Science Association Inc.

  15. miR-22 inhibits mouse ovarian granulosa cell apoptosis by targeting SIRT1.

    Science.gov (United States)

    Xiong, Fang; Hu, Lingqing; Zhang, Yun; Xiao, Xiao; Xiao, Juxia

    2016-02-24

    Granulosa cell (GC) apoptosis has been shown to be involved in follicular atresia, which is a degenerative process in ovarian follicles of mammals. However, the mechanism underlying the regulation of follicular atresia, particularly by microRNAs, is not well known. Real-time PCR (RT-PCR) was used to detect the expression level of miR-22 in healthy follicles (HF), early atretic follicles (EAF), and progressively atretic follicles (PAF). Flow cytometry was performed to assess the apoptosis of mouse granulosa cells (mGCs) treated with miR-22 mimics or negative control (NC) mimics. Regulation of the expression of SIRT1 by miR-22 was evaluated using a luciferase reporter assay system. To investigate the roles of SIRT1 in mGC apoptosis, the endogenous SIRT1 gene in mGCs was knocked down using an siRNA specific for SIRT1. miR-22 was increased during follicular atresia and suppressed granulosa cell apoptosis. The results of the luciferase reporter assay indicated that SIRT1 was a target gene of miR-22. In addition, knockdown of SIRT1 attenuated apoptosis in mGCs. miR-22 inhibits mGC apoptosis by downregulating SIRT1 directly in vitro. This study provides important insights into understanding the regulation mechanism of ovarian follicle atresia.

  16. Transcriptome profile of one-month-old lambs’ granulosa cells after superstimulation

    Science.gov (United States)

    Wu, Yangsheng; Lin, Jiapeng; Li, Xiaolin; Han, Bing; Wang, Liqin; Liu, Mingjun; Huang, Juncheng

    2017-01-01

    Objective Superstimulatory treatment of one-month-old lambs can achieve synchronous development of numerous growing follicles. However, these growing follicles cannot complete maturation and ovulation. Oocyte maturation and competence are acquired during follicular development, in which granulosa cells play an essential role. Methods In this study, we applied RNA sequencing to analyze and compare gene expression between prepubertal and adult superstimulated follicle granulosa cells in sheep. Results There were more than 300 genes that significantly differed in expression. Among these differently expressed genes, many extracellular matrix genes (EGF containing Fibulin Like Extracellular Matrix Protein 1, pentraxin 3, adrenomedullin, and osteopontin) were significantly down-regulated in the superstimulated follicles. Ingenuity pathway and gene ontology analyses revealed that processes of axonal guidance, cell proliferation and DNA replication were expressed at higher levels in the prepubertal follicles. Epidermal growth factor, T-Box protein 2 and beta-estradiol upstream regulator were predicted to be active in prepubertal follicles. By comparison, tumor protein P53 and let-7 were most active in adult follicles. Conclusion These results may contribute to a better understanding of the mechanisms governing the development of granulosa cells in the growing follicle in prepubertal sheep. PMID:27189640

  17. Effect of punicalagin on proliferation of porcine ovarian granulosa cells in vitro

    Directory of Open Access Journals (Sweden)

    Dagmara Packová

    2016-12-01

    Full Text Available Punicalagin is a major component responsible for pomegranate's (Punica granatum antioxidant properties. Punicalagin is the predominant ellagitannin of Punica granatum and present in two isomeric forms: punicalagin α and β. Punicalagin is metabolised to ellagic acid (antioxidant and microorganisms present in colon can metabolize ellagic acid to urolithins. The aim of in vitro study was to examine the effect of punicalagin on mitochondrial activity and markers of proliferation in porcine ovarian granulosa cells. The cells were cultivated during 24h without (control group and with various doses (0.01, 0.1, 1, 10 and 100 μg*ml-1 of pomegranate compound – punicalagin. MTT assay and immunocytochemistry were used in this study. Stimulatory influence of punicalagin on the mitochondrial activity of ovarian granulosa cells at concentrations 1 μg*ml-1 was found. Punicalagin (at 1 μg*ml-1 had a significant (P < 0.05 impact on the presence of proliferative markers cyclin B1 (increase and PCNA - proliferating cell nuclear antigen (decrease in porcine ovarian granulosa cells. These results suggest dose-dependent effect of punicalagin on cell proliferation. Further verification of possible role of punicalagin in proliferation is therefore needed.

  18. Management of Ocular Diseases Using Lutein and Zeaxanthin: What Have We Learned from Experimental Animal Studies?

    Science.gov (United States)

    Xue, Chunyan; Rosen, Richard; Jordan, Adrienne; Hu, Dan-Ning

    2015-01-01

    Zeaxanthin and lutein are two carotenoid pigments that concentrated in the retina, especially in the macula. The effects of lutein and zeaxanthin on the prevention and treatment of various eye diseases, including age-related macular degeneration, diabetic retinopathy and cataract, ischemic/hypoxia induced retinopathy, light damage of the retina, retinitis pigmentosa, retinal detachment, and uveitis, have been studied in different experimental animal models. In these animal models, lutein and zeaxanthin have been reported to have beneficial effects in protecting ocular tissues and cells (especially the retinal neurons) against damage caused by different etiological factors. The mechanisms responsible for these effects of lutein and zeaxanthin include prevention of phototoxic damage by absorption of blue light, reduction of oxidative stress through antioxidant activity and free radical scavenging, and their anti-inflammatory and antiangiogenic properties. The results of these experimental animal studies may provide new preventive and therapeutic procedures for clinical management of various vision-threatening diseases.

  19. Management of Ocular Diseases Using Lutein and Zeaxanthin: What Have We Learned from Experimental Animal Studies?

    Directory of Open Access Journals (Sweden)

    Chunyan Xue

    2015-01-01

    Full Text Available Zeaxanthin and lutein are two carotenoid pigments that concentrated in the retina, especially in the macula. The effects of lutein and zeaxanthin on the prevention and treatment of various eye diseases, including age-related macular degeneration, diabetic retinopathy and cataract, ischemic/hypoxia induced retinopathy, light damage of the retina, retinitis pigmentosa, retinal detachment, and uveitis, have been studied in different experimental animal models. In these animal models, lutein and zeaxanthin have been reported to have beneficial effects in protecting ocular tissues and cells (especially the retinal neurons against damage caused by different etiological factors. The mechanisms responsible for these effects of lutein and zeaxanthin include prevention of phototoxic damage by absorption of blue light, reduction of oxidative stress through antioxidant activity and free radical scavenging, and their anti-inflammatory and antiangiogenic properties. The results of these experimental animal studies may provide new preventive and therapeutic procedures for clinical management of various vision-threatening diseases.

  20. Inhibition of listeriolysin O oligomerization by lutein prevents Listeria monocytogenes infection.

    Science.gov (United States)

    Liu, Bowen; Teng, Zihao; Wang, Jianfeng; Lu, Gejin; Deng, Xuming; Li, Li

    2017-01-01

    The foodborne pathogenic bacterial species Listeria monocytogenes (L. monocytogenes) has caused incalculable damages to public health, and its successful infection requires various virulence factors, including Listeriolysin O (LLO). By forming pores in phagosomal membranes and even in some organelles, LLO plays an indispensable role in the ability of L. monocytogenes to escape from host immune attacks. Because of its critical role, LLO offers an appropriate therapeutic target against L. monocytogenes infection. Here, lutein, a natural small molecule existing widely in fruits and vegetables, is demonstrated as an effective inhibitor of LLO that works by blocking its oligomerization during invasion without showing significant bacteriostatic activity. Further assays applying lutein in cell culture models of invasion and in animal models showed that lutein could effectively inhibit L. monocytogenes infection. Overall, our results indicate that lutein may represent a promising and novel therapeutic agent against L. monocytogenes infection. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. Extraction of Lutein Diesters from Tagetes Erecta using Supercritical CO2 and Liquid Propane.

    Science.gov (United States)

    Skerget, Mojca; Bezjak, Miran; Makovšek, Katja; Knez, Zeljko

    2010-03-01

    The efficiency of high pressure extraction of lutein diesters from marigold (Tagetes erecta) flower petals has been investigated. The solvents used for extraction were supercritical carbon dioxide and liquid propane. Operating parameters were 300 bar and 40, 60 and 80 °C for CO2 and 100, 150, 200 bar and 40 and 60 °C for propane, respectively. The influence of process parameters on the total yield of extraction and content of lutein diesters in the extracts was investigated. The results show, that solvent power of propane for lutein diesters is approximately 3.5 times higher than of CO2. The calculation procedure based on the Fick's second law was applied to determine the diffusivities of lutein diesters during extraction from marigold flower petals for both extraction stages: a constant rate stage followed by a stage of decreasing rate. The mathematical model based on the Fick's second law well described the experimental extraction results.

  2. Intake of Lutein-Rich Vegetables Is Associated with Higher Levels of Physical Activity.

    Science.gov (United States)

    Crichton, Georgina; Elias, Merrill; Alkerwi, Ala'a; Buckley, Jonathon

    2015-09-18

    Levels of physical inactivity, a major contributor to burden of disease, are high in many countries. Some preliminary research suggests that circulating lutein concentrations are associated with high levels of physical activity (PA). We aimed to assess whether the intake of lutein-containing foods, including vegetables and eggs, is associated with levels of PA in two studies conducted in different countries. Dietary data and PA data collected from participants in two cross-sectional studies: the Maine-Syracuse Longitudinal Study (MSLS), conducted in Central New York, USA (n = 972), and the Observation of Cardiovascular Risk Factors in Luxembourg Study (ORISCAV-LUX) (n = 1331) were analyzed. Higher intakes of lutein containing foods, including green leafy vegetables, were associated with higher levels of PA in both study sites. Increasing the consumption of lutein-rich foods may have the potential to impact positively on levels of PA. This needs to be further explored in randomized controlled trials.

  3. Intake of Lutein-Rich Vegetables Is Associated with Higher Levels of Physical Activity

    Directory of Open Access Journals (Sweden)

    Georgina Crichton

    2015-09-01

    Full Text Available Levels of physical inactivity, a major contributor to burden of disease, are high in many countries. Some preliminary research suggests that circulating lutein concentrations are associated with high levels of physical activity (PA. We aimed to assess whether the intake of lutein-containing foods, including vegetables and eggs, is associated with levels of PA in two studies conducted in different countries. Dietary data and PA data collected from participants in two cross-sectional studies: the Maine-Syracuse Longitudinal Study (MSLS, conducted in Central New York, USA (n = 972, and the Observation of Cardiovascular Risk Factors in Luxembourg Study (ORISCAV-LUX (n = 1331 were analyzed. Higher intakes of lutein containing foods, including green leafy vegetables, were associated with higher levels of PA in both study sites. Increasing the consumption of lutein-rich foods may have the potential to impact positively on levels of PA. This needs to be further explored in randomized controlled trials.

  4. Oxidative Stress Induced by Zearalenone in Porcine Granulosa Cells and Its Rescue by Curcumin In Vitro

    Science.gov (United States)

    Qin, Xunsi; Cao, Mingjun; Lai, Fangnong; Yang, Fan; Ge, Wei; Zhang, Xifeng; Cheng, Shunfeng; Sun, Xiaofeng; Qin, Guoqing; Shen, Wei; Li, Lan

    2015-01-01

    Oxidative stress (OS), as a signal of aberrant intracellular mechanisms, plays key roles in maintaining homeostasis for organisms. The occurrence of OS due to the disorder of normal cellular redox balance indicates the overproduction of reactive oxygen species (ROS) and/or deficiency of antioxidants. Once the balance is broken down, repression of oxidative stress is one of the most effective ways to alleviate it. Ongoing studies provide remarkable evidence that oxidative stress is involved in reproductive toxicity induced by various stimuli, such as environmental toxicants and food toxicity. Zearalenone (ZEA), as a toxic compound existing in contaminated food products, is found to induce mycotoxicosis that has a significant impact on the reproduction of domestic animals, especially pigs. However, there is no information about how ROS and oxidative stress is involved in the influence of ZEA on porcine granulosa cells, or whether the stress can be rescued by curcumin. In this study, ZEA-induced effect on porcine granulosa cells was investigated at low concentrations (15 μM, 30 μM and 60 μM). In vitro ROS levels, the mRNA level and activity of superoxide dismutase, glutathione peroxidase and catalase were obtained. The results showed that in comparison with negative control, ZEA increased oxidative stress with higher ROS levels, reduced the expression and activity of antioxidative enzymes, increased the intensity of fluorogenic probes 2’, 7’-Dichlorodihydrofluorescin diacetate and dihydroethidium in flow cytometry assay and fluorescence microscopy. Meanwhile, the activity of glutathione (GSH) did not change obviously following 60 μM ZEA treatment. Furthermore, the underlying protective mechanisms of curcumin on the ZEA-treated porcine granulosa cells were investigated. The data revealed that curcumin pre-treatment significantly suppressed ZEA-induced oxidative stress. Collectively, porcine granulosa cells were sensitive to ZEA, which may induce oxidative

  5. Oxidative stress induced by zearalenone in porcine granulosa cells and its rescue by curcumin in vitro.

    Directory of Open Access Journals (Sweden)

    Xunsi Qin

    Full Text Available Oxidative stress (OS, as a signal of aberrant intracellular mechanisms, plays key roles in maintaining homeostasis for organisms. The occurrence of OS due to the disorder of normal cellular redox balance indicates the overproduction of reactive oxygen species (ROS and/or deficiency of antioxidants. Once the balance is broken down, repression of oxidative stress is one of the most effective ways to alleviate it. Ongoing studies provide remarkable evidence that oxidative stress is involved in reproductive toxicity induced by various stimuli, such as environmental toxicants and food toxicity. Zearalenone (ZEA, as a toxic compound existing in contaminated food products, is found to induce mycotoxicosis that has a significant impact on the reproduction of domestic animals, especially pigs. However, there is no information about how ROS and oxidative stress is involved in the influence of ZEA on porcine granulosa cells, or whether the stress can be rescued by curcumin. In this study, ZEA-induced effect on porcine granulosa cells was investigated at low concentrations (15 μM, 30 μM and 60 μM. In vitro ROS levels, the mRNA level and activity of superoxide dismutase, glutathione peroxidase and catalase were obtained. The results showed that in comparison with negative control, ZEA increased oxidative stress with higher ROS levels, reduced the expression and activity of antioxidative enzymes, increased the intensity of fluorogenic probes 2', 7'-Dichlorodihydrofluorescin diacetate and dihydroethidium in flow cytometry assay and fluorescence microscopy. Meanwhile, the activity of glutathione (GSH did not change obviously following 60 μM ZEA treatment. Furthermore, the underlying protective mechanisms of curcumin on the ZEA-treated porcine granulosa cells were investigated. The data revealed that curcumin pre-treatment significantly suppressed ZEA-induced oxidative stress. Collectively, porcine granulosa cells were sensitive to ZEA, which may induce

  6. Enhanced production of lutein in heterotrophic Chlorella protothecoides by oxidative stress

    Institute of Scientific and Technical Information of China (English)

    WEI Dong; CHEN Feng; CHEN Gu; ZHANG XueWu; LIU LongJun; ZHANG Hao

    2008-01-01

    The fast growing unicellular green microalgae Chlorella protothecoides has attracted interest as a promising organism for commercial production of a high-value carotenoid, lutein, by heterotrophic fermentation. Effects of two oxidant-forming reactive oxygen species (ROS) on the biomass concentration, and yield and content of lutein in batch culture of heterotrophic Chlorella protothecoides were investigated in this study. The addition of 0.1 mmol/L H2O2 and 0.01 mmol/L NaClO plus 0.5 mmol/L Fe2+ to the culture led to the generation of OH and enhanced the lutein content from 1.75 to 1.90 and 1.95 mg/g, respectively. The lutein content further increased to 1.98 mg/g when 0.01 mmol/L H2O2 and 0.5 mmol/L NaClO were added to generate 1O2. The maximum yield of lutein (28.5, 29.8 and 31.4 mg/L) and a high biomass concentration (15.0, 15.3 and 15.9 g/L) were also achieved through the above treatments. The results indicated that 1O2 could promote lutein formation and enhance lutein production in heterotrophic Chlorella protothecoides. Moreover, 1O2 produced from the reaction of H2O2 and NaClO was more effective in enhancing lutein production and reducing biomass loss than OH from the reaction of H2O2 or NaClO plus Fe2+.

  7. Enhanced production of lutein in heterotrophic Chlorella protothecoides by oxidative stress

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The fast growing unicellular green microalgae Chlorella protothecoides has attracted interest as a promising organism for commercial production of a high-value carotenoid, lutein, by heterotrophic fermentation. Effects of two oxidant-forming reactive oxygen species (ROS) on the biomass concen-tration, and yield and content of lutein in batch culture of heterotrophic Chlorella protothecoides were investigated in this study. The addition of 0.1 mmol/L H2O2 and 0.01 mmol/L NaClO plus 0.5 mmol/L Fe2+ to the culture led to the generation of ·OH and enhanced the lutein content from 1.75 to 1.90 and 1.95 mg/g, respectively. The lutein content further increased to 1.98 mg/g when 0.01 mmol/L H2O2 and 0.5 mmol/L NaClO were added to generate 1O2. The maximum yield of lutein (28.5, 29.8 and 31.4 mg/L) and a high biomass concentration (15.0, 15.3 and 15.9 g/L) were also achieved through the above treatments. The results indicated that 1O2 could promote lutein formation and enhance lutein production in hetero-trophic Chlorella protothecoides. Moreover, 1O2 produced from the reaction of H2O2 and NaClO was more effective in enhancing lutein production and reducing biomass loss than ·OH from the reaction of H2O2 or NaClO plus Fe2+.

  8. Macular Pigment and Lutein Supplementation in ABCA4-associated Retinal Degenerations

    Science.gov (United States)

    Aleman, Tomas S.; Cideciyan, Artur V.; Windsor, Elizabeth A. M.; Schwartz, Sharon B.; Swider, Malgorzata; Chico, John D.; Sumaroka, Alexander; Pantelyat, Alexander Y.; Duncan, Keith G.; Gardner, Leigh M.; Emmons, Jessica M.; Steinberg, Janet D.; Stone, Edwin M.; Jacobson, Samuel G.

    2008-01-01

    PURPOSE To determine macular pigment (MP) optical density (OD) in patients with ABCA4-associated retinal degenerations (ABCA4-RD) and the response of MP and vision to supplementation with lutein. METHODS Stargardt disease or cone-rod dystrophy patients with foveal fixation and with known or suspected disease-causing mutations in the ABCA4 gene were included. MPOD profiles were measured with heterochromatic flicker photometry. Serum carotenoids, visual acuity, foveal sensitivity and retinal thickness were quantified. Changes in MPOD and central vision were determined in a subset of patients receiving oral supplementation with lutein for 6 months. RESULTS MPOD in patients ranged from normal to markedly abnormal. As a group, ABCA4-RD patients had reduced foveal MPOD and there was strong correlation with retinal thickness. Average foveal tissue concentration of MP, estimated by dividing MPOD by retinal thickness, was normal in patients whereas serum concentration of lutein and zeaxanthin was significantly lower than normal. After oral lutein supplementation for 6 months, 91% of the patients showed significant increases in serum lutein and 63% of the patient eyes showed a significant augmentation in MPOD. The retinal responders tended to be female, and have lower serum lutein and zeaxanthin, lower MPOD and greater retinal thickness at baseline. Responding eyes had significantly lower baseline MP concentration compared to non-responding eyes. Central vision was unchanged after the period of supplementation. CONCLUSIONS MP is strongly affected by the stage of ABCA4 disease leading to abnormal foveal architecture. MP could be augmented by supplemental lutein in some patients. There was no change in central vision after 6 months of lutein supplementation. Long-term influences on the natural history of this supplement on macular degenerations require further study. PMID:17325179

  9. Solitary luteinized follicle cyst of pregnancy complicated with persistent postpartum vaginal bleeding: case report

    Institute of Scientific and Technical Information of China (English)

    ZHANG Song-ying; HUANG He-feng; TONG Xiao-mei

    2007-01-01

    @@ Solitary luteinized follicle cyst, a rare cause of ovarian enlargement during pregnancy and puerperium, is a self-limited disease that can regress spontaneously after labor. The complications of the disease include ovarian torsion, intracystic hemorrhage, and rupture; endocrine disturbances have not been reported.1-4 Here we report a case of solitary luteinized follicle cyst of pregnancy,which required surgical intervention owing to persistent postpartum vaginal bleeding.

  10. Expression of luteinizing hormone receptors in the mouse penis.

    Science.gov (United States)

    Kokk, Kersti; Kuuslahti, Marianne; Keisala, Tiina; Purmonen, Sami; Kaipia, Antti; Tammela, Teuvo; Orro, Helen; Simovart, Helle-Evi; Pöllänen, Pasi

    2011-01-01

    The role of luteinizing hormone (LH) in the regulation of normal reproductive functions in males and females is quite well established. Besides the expression of LH receptors in the target cells in gonads, it has been found in several extragonadal organs. There is no information about the expression of LH receptors in the penis up to now. The aim of the present study is to investigate the expression of the LH receptor in the mouse penis to see if LH effects are possible in the penis. BALB/c mice were used as donors of normal penis and testis tissue. Immunocytochemistry, Western blotting, and quantitative reverse transcriptase polymerase chain reactions (RT-PCRs) were used for the detection of the LH receptor. Positive immunoreaction for LH receptors was present in the nuclei of urethral epithelium and endothelial cells of cavernous spaces in the corpus cavernosum and corpus spongiosum penis. Western blotting experiments demonstrated the presence of LH antigen at M(r) = 97.4 and 78 kd. Quantitative RT-PCRs confirmed the expression of LH receptor in the penis. Our results show that LH receptor is expressed in the body of the mouse penis; thus, it may directly regulate functions of penile tissue.

  11. The Relationship between Lutein and Zeaxanthin Status and Body Fat

    Directory of Open Access Journals (Sweden)

    Billy R. Hammond

    2013-03-01

    Full Text Available The objective of this project was to investigate the relationships between total and regional distribution of body fat and tissue lutein (L and zeaxanthin (Z status. Healthy men and women (N = 100; average age: 22.5 year, average BMI: 23.4 kg/m2 were evaluated. Total body and regional fat mass were assessed by dual-energy X-ray absorptiometry (Hologic Delphi A. Serum LZ was measured using reverse phase high-performance liquid chromatography, and retinal LZ (referred to as macular pigment optical density; MPOD was measured using heterochromatic flicker photometry. Body fat percentage (total and regional was inversely related to MPOD (p < 0.01 but no significant relationship was found for serum LZ. Higher body fat percentage, even within relatively healthy limits, is associated with lower tissue LZ status. The results indicate that adiposity may affect the nutritional state of the retina. Such links may be one of the reasons that obesity promotes age-related degenerative conditions of the retina.

  12. Lutein epoxide cycle, more than just a forest tale

    Science.gov (United States)

    Becerril, José María; García-Plazaola, José Ignacio

    2009-01-01

    Two xanthophyll cycles have been described in higher plants: the ubiquitous violaxanthin (V) cycle and the taxonomically restricted lutein epoxide (Lx) cycle. Both involve the light induced de-epoxidation of an epoxidated xanthophyll (V or Lx) and the epoxidation back in the dark. Evolutionary trends and function of the Lx cycle are still not clear. Up to nowadays, significant amounts of Lx have been found in several unrelated taxa, but it is a character almost exclusive from woody plants (except in the case of the parasitic plant Cuscuta reflexa). We have found an exception to this pattern in Cucumis sativus L., which showed high concentrations of Lx. Since Lx cycle was operative in leaves and cotyledons of this species and Lx concentration were much higher in cotyledons than in leaves, we speculate a role for the early stages of development. To date, this species is the first herbaceous non-parasitic species with operative Lx cycle. Since this species can be much more easily and rapidly grown and investigated than woody plants, these data can open new horizons and new lines of investigation for Lx cycle. PMID:19794858

  13. Morphological and Hormonal Identiifcation of Porcine Atretic Follicles and Relationship Analysis of Hormone Receptor Levels During Granulosa Cell Apoptosis In vivo

    Institute of Scientific and Technical Information of China (English)

    YU De-bing; YU Min-li; LIN Fei; JIANG Bao-chun; YANG Li-na; WANG Si-yu; ZHAO Ying; WNAG Zheng-chao

    2014-01-01

    Recent reports have demonstrated that follicular atresia is initiated or caused by granulosa cell apoptosis followed by theca cell degeneration in mammalian ovaries, but the mechanism of follicular atresia is still to be elucidated. Therefore, our present study was designed to examine our hypothesis that the changes of follicular microenvironment induce the granulosa cell apoptosis during pocrine follicular atresia in vivo. We ifrstly isolated intact porcine antral follicles and identiifed them into three groups, healthy follicles (HF), early atretic follicles (EAF) and progressed atretic follicles (PAF) through morphology and histology. To further conifrm their status, we detected hormone levels in follicular lfuids and the expression level of apoptosis gene Bax in granulosa cells. The rate of progesterone (P) and estradiol (E2) was increased with the expression of Bax, indicating hormone can be used as a marker of granulosa cell apoptosis or follicular atresia. Finally, we analyzed the expression level of hormone receptor genes in granulosa cells and their relationship with follicular atresia. In PAF, the expression of Progesterone receptor (PGR) was increased signiifcantly while estradiol receptor (ER) had no notable changes, which suggesting the increased-PGR accelerated the effect of P-stimulated granulosa cell apoptosis. The dramatic increasing of androgen receptor (AR) expression in PAF and the obvious increase of tumor necrosis factor-αreceptor (TNFR) in EAF indicated that there are different pathways regulating granulosa cell apoptosis during follicular atresia. Together, our results suggested that different pathways of granulosa cell apoptosis was induced by changing the follicular microenvironment during follicular atresia.

  14. Follicle-stimulating Hormone (FSH) Induced Internalization of Porcine FSH Receptor in Cultured Porcine Granulosa Cells and Chinese Hamster Ovary Cells Transfected with Recombinant Porcine FSH Receptor cDNA

    Institute of Scientific and Technical Information of China (English)

    ZHU; Changhong; TIAN; Hong; XIONG; Zhongming; XIA; Huizhu

    2001-01-01

    In order to study the fate of human follicle-stimulating hormone (FSH) when hormone binds to its receptor, a quick biochemical method that can differentiate between the surface-bound and internalized hormone was used to determine the internalization induced by FSH in cultured both porcine granulosa cells and Chinese hamster ovary (CHO) cells expressing recombinant porcine FSH receptor. The results showed that FSH was slowly internalized, and the internalized radioactivity (acid resistant) reached a peak 10-12 h after addition of 125I-hFSH. It was suggested that FSHR do not get internalized rapidly under physiological circumstances precisely because the appropriate sequences are absent.

  15. The Pharmacological Effects of Lutein and Zeaxanthin on Visual Disorders and Cognition Diseases.

    Science.gov (United States)

    Jia, Yu-Ping; Sun, Lei; Yu, He-Shui; Liang, Li-Peng; Li, Wei; Ding, Hui; Song, Xin-Bo; Zhang, Li-Juan

    2017-04-20

    Lutein (L) and zeaxanthin (Z) are dietary carotenoids derived from dark green leafy vegetables, orange and yellow fruits that form the macular pigment of the human eyes. It was hypothesized that they protect against visual disorders and cognition diseases, such as age-related macular degeneration (AMD), age-related cataract (ARC), cognition diseases, ischemic/hypoxia induced retinopathy, light damage of the retina, retinitis pigmentosa, retinal detachment, uveitis and diabetic retinopathy. The mechanism by which they are involved in the prevention of eye diseases may be due their physical blue light filtration properties and local antioxidant activity. In addition to their protective roles against light-induced oxidative damage, there are increasing evidences that L and Z may also improve normal ocular function by enhancing contrast sensitivity and by reducing glare disability. Surveys about L and Z supplementation have indicated that moderate intakes of L and Z are associated with decreased AMD risk and less visual impairment. Furthermore, this review discusses the appropriate consumption quantities, the consumption safety of L, side effects and future research directions.

  16. The Pharmacological Effects of Lutein and Zeaxanthin on Visual Disorders and Cognition Diseases

    Directory of Open Access Journals (Sweden)

    Yu-Ping Jia

    2017-04-01

    Full Text Available Lutein (L and zeaxanthin (Z are dietary carotenoids derived from dark green leafy vegetables, orange and yellow fruits that form the macular pigment of the human eyes. It was hypothesized that they protect against visual disorders and cognition diseases, such as age-related macular degeneration (AMD, age-related cataract (ARC, cognition diseases, ischemic/hypoxia induced retinopathy, light damage of the retina, retinitis pigmentosa, retinal detachment, uveitis and diabetic retinopathy. The mechanism by which they are involved in the prevention of eye diseases may be due their physical blue light filtration properties and local antioxidant activity. In addition to their protective roles against light-induced oxidative damage, there are increasing evidences that L and Z may also improve normal ocular function by enhancing contrast sensitivity and by reducing glare disability. Surveys about L and Z supplementation have indicated that moderate intakes of L and Z are associated with decreased AMD risk and less visual impairment. Furthermore, this review discusses the appropriate consumption quantities, the consumption safety of L, side effects and future research directions.

  17. Effects of granulosa cells on steroidogenesis, proliferation and apoptosis of stromal cells and theca cells derived from the goat ovary.

    Science.gov (United States)

    Qiu, Mingning; Quan, Fusheng; Han, Chengquan; Wu, Bin; Liu, Jun; Yang, Zhongcai; Su, Feng; Zhang, Yong

    2013-11-01

    The aim of this study was to investigate the effect of granulosa cells from small antral follicles on steroidogenesis, proliferation and apoptosis of goat ovarian stromal and theca cells in vitro. Using Transwell co-culture system, we evaluated androgen production, LH responsiveness, cell proliferation and apoptosis and some molecular expression regarding steroidogenic enzyme and apoptosis-related genes in stromal and theca cells. The results indicated that the co-culture with granulosa cells increased steroidogenesis, LH responsiveness and bcl-2 gene expression as well as decreased apoptotic bax and bad expressions in stromal and theca cells. Thus, granulosa cells had a capacity of promoting steroidogenesis in stromal cell and LH responsiveness in cortical stromal cells, maintaining steroidogenesis in theca cells, inhibiting apoptosis of cortical stromal cells and improving anti-apoptotic abilities of stromal and theca cells.

  18. Effect of Carotenoid Supplemented Formula on Carotenoid Bioaccumulation in Tissues of Infant Rhesus Macaques: A Pilot Study Focused on Lutein

    Science.gov (United States)

    Jeon, Sookyoung; Neuringer, Martha; Johnson, Emily E.; Kuchan, Matthew J.; Pereira, Suzette L.; Johnson, Elizabeth J.; Erdman, John W.

    2017-01-01

    Lutein is the predominant carotenoid in the developing primate brain and retina, and may have important functional roles. However, its bioaccumulation pattern during early development is not understood. In this pilot study, we investigated whether carotenoid supplementation of infant formula enhanced lutein tissue deposition in infant rhesus macaques. Monkeys were initially breastfed; from 1 to 3 months of age they were fed either a formula supplemented with lutein, zeaxanthin, β-carotene and lycopene, or a control formula with low levels of these carotenoids, for 4 months (n = 2/group). All samples were analyzed by high pressure liquid chromatography (HPLC). Final serum lutein in the supplemented group was 5 times higher than in the unsupplemented group. All brain regions examined showed a selective increase in lutein deposition in the supplemented infants. Lutein differentially accumulated across brain regions, with highest amounts in occipital cortex in both groups. β-carotene accumulated, but zeaxanthin and lycopene were undetectable in any brain region. Supplemented infants had higher lutein concentrations in peripheral retina but not in macular retina. Among adipose sites, abdominal subcutaneous adipose tissue exhibited the highest lutein level and was 3-fold higher in the supplemented infants. The supplemented formula enhanced carotenoid deposition in several other tissues. In rhesus infants, increased intake of carotenoids from formula enhanced their deposition in serum and numerous tissues and selectively increased lutein in multiple brain regions. PMID:28075370

  19. Effect of Carotenoid Supplemented Formula on Carotenoid Bioaccumulation in Tissues of Infant Rhesus Macaques: A Pilot Study Focused on Lutein

    Directory of Open Access Journals (Sweden)

    Sookyoung Jeon

    2017-01-01

    Full Text Available Lutein is the predominant carotenoid in the developing primate brain and retina, and may have important functional roles. However, its bioaccumulation pattern during early development is not understood. In this pilot study, we investigated whether carotenoid supplementation of infant formula enhanced lutein tissue deposition in infant rhesus macaques. Monkeys were initially breastfed; from 1 to 3 months of age they were fed either a formula supplemented with lutein, zeaxanthin, β-carotene and lycopene, or a control formula with low levels of these carotenoids, for 4 months (n = 2/group. All samples were analyzed by high pressure liquid chromatography (HPLC. Final serum lutein in the supplemented group was 5 times higher than in the unsupplemented group. All brain regions examined showed a selective increase in lutein deposition in the supplemented infants. Lutein differentially accumulated across brain regions, with highest amounts in occipital cortex in both groups. β-carotene accumulated, but zeaxanthin and lycopene were undetectable in any brain region. Supplemented infants had higher lutein concentrations in peripheral retina but not in macular retina. Among adipose sites, abdominal subcutaneous adipose tissue exhibited the highest lutein level and was 3-fold higher in the supplemented infants. The supplemented formula enhanced carotenoid deposition in several other tissues. In rhesus infants, increased intake of carotenoids from formula enhanced their deposition in serum and numerous tissues and selectively increased lutein in multiple brain regions.

  20. Androstenedione increases cytochrome P450 aromatase messenger ribonucleic acid transcripts in nonluteinizing bovine granulosa cells.

    Science.gov (United States)

    Hamel, Mélanie; Vanselow, Jens; Nicola, Edmir S; Price, Christopher A

    2005-02-01

    The objective of this study was to determine if androgens regulate granulosa cell steroidogenesis at physiological doses found in small bovine follicles. Bovine granulosa cells were cultured under serum-free conditions that permit the induction and maintenance of FSH-dependent estradiol secretion. Increasing androstenedione concentrations from 0.1 to 1 or 10 microM significantly increased estradiol accumulation and cytochrome P450 aromatase (P450arom) mRNA abundance. No increase in progesterone accumulation or abundance of mRNA for P450 side-chain cleavage or 3beta-hydroxysteroid dehydrogenase enzymes was observed. The addition of 0.1, 1, or 10 microM progestins or estrogens had no stimulatory effect on P450arom mRNA levels. An analysis of the 5'-untranslated region of P450arom mRNA transcripts indicated that the majority was derived from Cyp19 ovary-specific promoter 2, with some contribution from promoters 1.1 and 1.5. Transcripts from these three promoters were all significantly increased by androstenedione. Testosterone increased promoter 1.1 and 1.5-derived transcripts, but only promoter 2-derived transcripts at the highest dose tested (100 microM). Dihydrotestosterone (DHT) did not affect Cyp19 expression. Collectively, these data show that androgens may exert specific stimulatory effects on P450arom mRNA concentrations in granulosa cells. Interestingly, different androgens had different effects on Cyp19 promoter usage, suggesting differential regulation of aromatase gene expression in the developing follicle.

  1. GnRH agonist versus GnRH antagonist in IVF/ICSI cycles with recombinant LH supplementation: DNA fragmentation and apoptosis in granulosa cells.

    Science.gov (United States)

    Lavorato, Heloisa L; Oliveira, Joao Batista A; Petersen, Claudia G; Vagnini, Laura; Mauri, Ana L; Cavagna, Mario; Baruffi, Ricardo L R; Franco, Jose G

    2012-11-01

    To compare the level of apoptosis and DNA fragmentation in the human granulosa cell (GC) layer exposed to an agonist or antagonist of GnRH in intracytoplasmic sperm injection (ICSI) cycles supplemented with recombinant LH (rLH). Patients without ovulatory dysfunction, aged ≤37 years and in their first ICSI cycle were prospectively randomised to receive either a long GnRH agonist protocol or a multi-dose antagonist protocol. In both groups, recombinant FSH supplemented with rLH was used for ovarian stimulation, and the GCs were collected during oocyte denudation. The GCs were then analysed for DNA fragmentation by TUNEL assay and for apoptosis using the annexin-V assay. The outcomes were given as the percentage of GCs with DNA fragmentation and apoptosis out of the total number of GCs analysed. Comparison of the agonist versus the antagonist group was performed using the Mann-Whitney test. DNA fragmentation: 32 patients were included in either the GnRH agonist group (n=16) or the antagonist group (n=16). The percentage of GCs with positive DNA fragmentation did not differ significantly (P=0.76) between the agonist group (15.5 ± 9.4%) and the antagonist group (18.8 ± 13.3%). Apoptosis: 28 patients were included in either the GnRH agonist group (n=14) or the antagonist group (n=14). The percentage of GCs positive for apoptosis did not differ significantly (P=0.78) between the agonist group (34.6 ± 14.7%) and the antagonist group (36.5 ± 22%). The results suggest that therapy with either an agonist or antagonist of GnRH is associated with comparable levels of DNA fragmentation and apoptosis in granulosa cells in ICSI cycles supplemented with rLH. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  2. A quantitative electron microscopic analysis of the membrana granulosa of rat preovulatory follicles.

    Science.gov (United States)

    Zoller, L C

    1984-01-01

    The ultrastructure of the membrana granulosa (MG) of rat preovulatory follicles was examined using stereological techniques. Organelles studied were nuclei, mitochondria, lipid droplets (LD), lysosomes, and smooth and rough endoplasmic reticulum (SER, RER). The peripheral region of the MG contained the greatest volume of mitochondria, LD and SER, organelles associated with steroidogenesis. The volume of RER, an organelle associated with protein production, was greatest in the cumulus oophorus. These results corroborate previous analyses and demonstrate that the rat MG is composed of discrete subregions.

  3. Mutual regulation of growth hormone and bone morphogenetic protein system in steroidogenesis by rat granulosa cells.

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    Nakamura, Eri; Otsuka, Fumio; Inagaki, Kenichi; Miyoshi, Tomoko; Matsumoto, Yoshinori; Ogura, Kanako; Tsukamoto, Naoko; Takeda, Masaya; Makino, Hirofumi

    2012-01-01

    GH induces preantral follicle growth and differentiation with oocyte maturation. However, the effects of GH on ovarian steroidogenesis and the mechanisms underlying its effects have yet to be elucidated. In this study, we investigated the actions of GH on steroidogenesis by rat granulosa cells isolated from early antral follicles by focusing on the ovarian bone morphogenetic protein (BMP) system. We found that GH suppressed FSH-induced estradiol production with reduction in aromatase expression and, in contrast, GH increased FSH-induced progesterone level with induction of steroidogenic acute regulatory protein, side chain cleavage cytochrome P450, and 3β-hydroxysteroid dehydrogenase. The effects of GH on steroidogenesis by granulosa cells were enhanced in the presence of the BMP antagonist noggin. Coculture of GH with oocytes did not alter GH regulation of steroidogenesis. Steroid production induced by cAMP donors was not affected by GH treatment and the GH effects on FSH-induced steroid production were not accompanied by changes in cAMP synthesis, suggesting that GH actions were not directly mediated by the cAMP-protein kinase A pathway. GH exerted synergistic effects on MAPK activation elicited by FSH, which regulated FSH-induced steroidogenesis. In addition, GH-induced signal transducer and activator of transcription phosphorylation was involved in the induction of IGF-I expression. GH increased IGF-I, IGF-I receptor, and FSH receptor expression in granulosa cells, and inhibition of IGF-I signaling restored GH stimulation of FSH-induced progesterone production, suggesting that endogenous IGF-I is functionally involved in GH effects on progesterone induction. BMP inhibited IGF-I effects that increased FSH-induced estradiol production with suppression of expression of the GH/IGF-I system, whereas GH/IGF-I actions impaired BMP-Sma and Mad related protein 1/5/8 signaling through down-regulation of the expression of BMP receptors. Thus, GH acts to modulate estrogen

  4. Parahippocampal Cortex Mediates the Relationship between Lutein and Crystallized Intelligence in Healthy, Older Adults

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    Marta Karolina Zamroziewicz

    2016-12-01

    Full Text Available Introduction: Although diet has a substantial influence on the aging brain, the relationship between dietary nutrients and aspects of brain health remains unclear. This study examines the neural mechanisms that mediate the relationship between a carotenoid important for brain health across the lifespan, lutein, and crystallized intelligence in cognitively intact older adults. We hypothesized that higher serum levels of lutein are associated with better performance on a task of crystallized intelligence, and that this relationship is mediated by gray matter structure of regions within the temporal cortex. This investigation aims to contribute to a growing line of evidence, which suggests that particular nutrients may slow or prevent aspects of cognitive decline by targeting specific features of brain aging.Methods: We examined 75 cognitively intact adults between the ages of 65 and 75 to investigate the relationship between serum lutein, tests of crystallized intelligence (measured by the Wechsler Abbreviated Scale of Intelligence, and gray matter volume of regions within the temporal cortex. A three-step mediation analysis was implemented using multivariate linear regressions to control for age, sex, education, income, depression status, and body mass index.Results: The mediation analysis revealed that gray matter thickness of one region within the temporal cortex, the right parahippocampal cortex (Brodmann’s Area 34, partially mediates the relationship between serum lutein and crystallized intelligence. Conclusion: These results suggest that the parahippocampal cortex acts as a mediator of the relationship between serum lutein and crystallized intelligence in cognitively intact older adults. Prior findings substantiate the individual relationships reported within the mediation, specifically the links between (i serum lutein and temporal cortex structure, (ii serum lutein and crystallized intelligence, and (iii parahippocampal cortex structure

  5. Safety assessment of lutein and zeaxanthin (Lutemax 2020): subchronic toxicity and mutagenicity studies.

    Science.gov (United States)

    Ravikrishnan, R; Rusia, Shraddha; Ilamurugan, G; Salunkhe, Ulhas; Deshpande, Jayant; Shankaranarayanan, J; Shankaranarayana, M L; Soni, Madhu G

    2011-11-01

    Lutein and zeaxanthin, naturally occurring carotenoids, have shown to reduce the risk of cataracts and age-related macular degeneration. Lutemax 2020 is a lutein and zeaxanthin (including meso-isomer) enriched product obtained from Marigold flowers (Tagetes erecta L). The objective of the present study was to investigate adverse effects, if any, of Lutemax 2020 in acute and subchronic toxicity, and mutagenicity studies. In acute toxicity study in rats no lethality was noted at 2000 mg Lutemax 2020/kg body weight (bw). In the subchronic study, Wistar rats (10/sex/group) were administered (gavage) lutein/zeaxanthin concentrate at dose levels of 0, 4, 40 and 400mg/kg bw/day for 90-days. Compared with the control group, administration of lutein/zeaxanthin concentrate did not result in any toxicologically significant treatment-related changes in clinical observations, ophthalmic examinations, body weights, body weight gains, feed consumption, and organ weights. No toxicologically relevant findings were noted in urinalysis, hematology or clinical biochemistry parameters at the end of the treatment or recovery period. Terminal necropsy did not reveal any treatment-related gross or histopathology findings. The results of mutagenicity testing in Salmonella typhimurium did not reveal any genotoxicity. The no observed-adverse-effect level (NOAEL) for lutein/zeaxanthin concentrate was determined as 400mg/kg bw/day, the highest dose tested. Copyright © 2011 Elsevier Ltd. All rights reserved.

  6. Micellarization and intestinal cell uptake of beta-carotene and lutein from drumstick (Moringa oleifera) leaves.

    Science.gov (United States)

    Pullakhandam, Raghu; Failla, Mark L

    2007-06-01

    The leaves and pods of the drumstick tree are used as food and medicine in some Asian and African countries. Although relatively high concentrations of beta-carotene and lutein have been reported in the leaves, the bioavailability of these carotenoids from this source is unknown. We have analyzed the digestive stability and bioaccessibility of carotenoids in fresh and lyophilized drumstick leaves using the coupled in vitro digestion/Caco-2 cell model. Beta-carotene and lutein were stable during simulated gastric and small intestinal digestion. The efficiency of micellarization of lutein during the small intestinal phase of digestion exceeded that of beta-carotene. Addition of peanut oil (5% vol/wt) to the test food increased micellarization of both carotenoids, and particularly beta-carotene. Caco-2 cells accumulated beta-carotene and lutein from micelles generated during digestion of drumstick leaves in a time- and concentration-dependent manner. The relatively high bioaccessibility of beta-carotene and lutein from drumstick leaves ingested with oil supports the potential use of this plant food for improving vitamin A nutrition and perhaps delaying the onset of some degenerative diseases such as cataracts.

  7. Outdoor cultivation of lutein-rich cells of Muriellopsis sp. in open ponds.

    Science.gov (United States)

    Blanco, Antonio M; Moreno, José; Del Campo, José A; Rivas, Joaquín; Guerrero, Miguel G

    2007-01-01

    The growth performance of the chlorophycean microalga Muriellopsis sp. outdoors in open tanks agitated with a paddlewheel and its ability to accumulate carotenoids have been evaluated throughout the year. The cells grown in the open system had free lutein as the main carotenoid, with violaxanthin, beta-carotene, and neoxanthin also present. Lutein content of the dry biomass ranged from 0.4 to 0.6%, depending on the growth and environmental conditions. In addition, the biomass of Muriellopsis sp. had a high content in both protein and lipids with about half of the fatty acids being of the polyunsaturated type, with alpha-linolenic acid accounting for almost 30% of the total fatty acids. The effect of determinant parameters on the performance of the cultures in open tanks was evaluated. Operating conditions that allow the maintenance of productive cultures were established under semicontinuous regime for 9 months throughout the year. Biomass and lutein yields in the open system were not far from those in closed tubular photobioreactors, and reached productivity values of 20 g dry biomass, containing around 100 mg lutein m(-2) day(-1) in summer. The outdoor culture of Muriellopsis sp. in open ponds thus represents a real alternative to established systems for the production of lutein.

  8. Characterization of luteinizing hormone and luteinizing hormone receptor and their indispensable role in the ovulatory process of the medaka.

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    Katsueki Ogiwara

    Full Text Available The molecular properties and roles of luteinizing hormone (Lh and its receptor (Lhcgrbb have not been studied for the medaka (Oryzias latipes, which is an excellent animal model for ovulation studies. Here, we characterized the medaka Lh/Lhcgrbb system, with attention to its involvement in the ovulatory process of this teleost fish. In the medaka ovary, follicle-stimulating hormone receptor mRNA was expressed in small and medium-sized follicles, while lhcgrbb mRNA was expressed in the follicle layers of all growing follicles. Experiments using HEK 293T cells expressing medaka Lhcgrbb in vitro revealed that gonadotropin from pregnant mare's serum and medaka recombinant Lh (rLh bound to the fish Lhcgrbb. The fish gonadotropin subunits Gtha, Fshb, and Lhb were essentially expressed at fairly constant levels in the pituitary of the fish during a 24-h spawning cycle. Using medaka rLh, we developed a follicle culture system that allowed us to follow the whole process of oocyte maturation and ovulation in vitro. This follicle culture method enabled us to determine that the Lh surge for the preovulatory follicle occurred in vivo between 19 and 15 h before ovulation. The present study also showed that oocyte maturation and ovulation were delayed several hours in vitro compared with in vivo. Treatment of large follicles with medaka rLh in vitro significantly increased the expression of Mmp15, which was previously demonstrated to be crucial for ovulation in the fish. These findings demonstrate that Lh/Lhcgrbb is critically involved in the induction of oocyte maturation and ovulation.

  9. Biodegradable Poly (Lactic-co-Glycolic Acid)-Polyethylene Glycol Nanocapsules: An Efficient Carrier for Improved Solubility, Bioavailability, and Anticancer Property of Lutein.

    Science.gov (United States)

    Arunkumar, Ranganathan; Prashanth, Keelara Veerappa Harish; Manabe, Yuki; Hirata, Takashi; Sugawara, Tatsuya; Dharmesh, Shylaja Mallaiah; Baskaran, Vallikannan

    2015-06-01

    Lutein bioavailability is limited because of its poor aqueous solubility. In this study, lutein-poly (lactic-co-glycolic acid) (PLGA)-polyethylene glycol (PEG) nanocapsules were prepared to improve the solubility, bioavailability, and anticancer property of lutein. The scanning electron microscopy and dynamic light scattering examination revealed that the nanocapsules are smooth and spherical with size ranging from 80 to 500 nm (mean = 200 nm). In vitro lutein release profile from nanocapsules showed controlled sustainable release (66%) up to 72 h. Aqueous solubility of lutein nanocapsules was much higher by 735-fold than the lutein. Fourier transform infrared spectroscopy analyses showed no chemical interaction among PLGA, PEG, and lutein, indicating possible weak intermolecular forces like hydrogen bonds. X-ray diffraction revealed lutein is distributed in a disordered amorphous state in nanocapsules. Postprandial plasma kinetics (area under the curve) of an oral dose of lutein from nanocapsules was higher by 5.4-fold compared with that of micellar lutein (control). The antiproliferative effect of lutein from nanocapsules (IC50 value, 10.9 μM) was higher (43.6%) than the lutein (IC50 value, 25 μM). Results suggest that PLGA-PEG nanocapsule is an efficient carrier for enhancing hydrophilicity, bioavailability, and anticancer property of lipophilic molecules such as lutein.

  10. Clinicopathologic Analysis of Ovary Guvenile Granulosa Cell Tumor%卵巢幼年型粒层细胞瘤临床病理观察

    Institute of Scientific and Technical Information of China (English)

    崔华娟; 赖日权; 王卓才; 王炜; 彭大云

    2012-01-01

    目的 探讨卵巢幼年型粒层细胞瘤(juvenile granulosa cell tumor,JGCT)的临床病理特征、早期诊断与预后的关系.方法 报道2例少见的幼年型卵巢粒层细胞瘤,并结合国内外文献进行讨论.结果 2例均为青年女性,分别为21岁和27岁.例1为肩胛骨软骨肉瘤术后3年出现腹胀,术后诊断为JGCT,FIGOⅢ期,随访26个月死亡.例2表现为月经失调,临床诊断为FIGOⅠA期,术后随访29个月未见复发.镜下全部为弥漫生长和不典型滤泡样结构,未见Call-Exner小体.肿瘤细胞核小、较圆、深染,极少见核沟,例1细胞重度异型,核分裂象>10/10 HPF;例2细胞轻度异型,核分裂象<5/10 HPF.免疫组化2例vimentin均强阳性,CD99散在弱阳性;例1滤泡内物质黏液卡红染色阳性,例2 α抑制素(α-inhibin)小灶阳性;CK18阳性,其余一抗细胞角蛋白(cytokeratin pan,CKpan)、S-100蛋白、蛋白基因产物(protein gene product,PGP)9.5、平滑肌肌动蛋白(smooth muscle actin,SMA)、上皮膜抗原(epithelial membrane antigen,EMA)、肌源性标记物结蛋白(desmin)、肌细胞生成素(myogenin)、血管内皮细胞生长因子(vascular endothelial growth factor,VEGF)、胎盘碱性磷酸酶(placental alkaline phosphatase,PLAP)、人体绒毛膜促性腺激素(human chorionic gonagotropin,HCG)、甲胎蛋白(alpha-fetoprotein,AFP)、CD45、Calretinin均阴性.结论 JGCT属于交界性或低度恶性肿瘤,但少数病例恶性程度较高,早期诊断是预后的重要因素,需要与卵巢成年型粒层细胞瘤、高血钙型小细胞癌、恶性黑色素瘤等进行鉴别.%Objective To study the relationship among the clinicopathologic features, early diagnose and prognosis of ovary juvenile granulosa cell tumor. Methods Two cases with ovary juvenile granulosa cell tumor were reported and the correlative literature was reviewed. Results The two cases were young females, 21 and 27 years old respectively. One case presented with swelling

  11. Human LH and hCG stimulate differently the early signalling pathways but result in equal testosterone synthesis in mouse Leydig cells in vitro.

    Science.gov (United States)

    Riccetti, Laura; De Pascali, Francesco; Gilioli, Lisa; Potì, Francesco; Giva, Lavinia Beatrice; Marino, Marco; Tagliavini, Simonetta; Trenti, Tommaso; Fanelli, Flaminia; Mezzullo, Marco; Pagotto, Uberto; Simoni, Manuela; Casarini, Livio

    2017-01-05

    Human luteinizing hormone (LH) and chorionic gonadotropin (hCG) are glycoprotein hormones regulating development and reproductive functions by acting on the same receptor (LHCGR). We compared the LH and hCG activity in gonadal cells from male mouse in vitro, i.e. primary Leydig cells, which is a common tool used for gonadotropin bioassay. Murine Leydig cells are naturally expressing the murine LH receptor (mLhr), which binds human LH/hCG. Cultured Leydig cells were treated by increasing doses of recombinant LH and hCG, and cell signaling, gene expression and steroid synthesis were evaluated. We found that hCG is about 10-fold more potent than LH in cAMP recruitment, and slightly but significantly more potent on cAMP-dependent Erk1/2 phosphorylation. However, no significant differences occur between LH and hCG treatments, measured as activation of downstream signals, such as Creb phosphorylation, Stard1 gene expression and testosterone synthesis. These data demonstrate that the responses to human LH/hCG are only quantitatively and not qualitatively different in murine cells, at least in terms of cAMP and Erk1/2 activation, and equal in activating downstream steroidogenic events. This is at odds with what we previously described in human primary granulosa cells, where LHCGR mediates a different pattern of signaling cascades, depending on the natural ligand. This finding is relevant for gonadotropin quantification used in the official pharmacopoeia, which are based on murine, in vivo bioassay and rely on the evaluation of long-term, testosterone-dependent effects mediated by rodent receptor.

  12. Rhesus monkey cumulus cells revert to a mural granulosa cell state after an ovulatory stimulus.

    Science.gov (United States)

    Chaffin, Charles L; Lee, Young S; VandeVoort, Catherine A; Patel, Bela G; Latham, Keith E

    2012-11-01

    Follicular somatic cells (mural granulosa cells and cumulus cells) and the oocyte communicate through paracrine interactions and through direct gap junctions between oocyte and cumulus cells. Considering that mural and cumulus cells arise through a common developmental pathway and that their differentiation is essential to reproductive success, understanding how these cells differ is a key aspect to understanding their critical functions. Changes in global gene expression before and after an ovulatory stimulus were compared between cumulus and mural granulosa cells to test the hypothesis that mural and cumulus cells are highly differentiated at the time of an ovulatory stimulus and further differentiate during the periovulatory interval. The transcriptomes of the two cell types were markedly different (>1500 genes) before an ovulatory hCG bolus but converged after ovulation to become completely overlapping. The predominant transition was for the cumulus cells to become more like mural cells after hCG. This indicates that the differentiated phenotype of the cumulus cell is not stable and irreversibly established but may rather be an ongoing physiological response to the oocyte.

  13. Central precocious puberty and granulosa cell ovarian tumor in an 8-year old female

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    Valeria Calcaterra

    2013-07-01

    Full Text Available Ovarian tumors associated with hormonal changes of the peripheral iso-sexual precocious puberty are of common presentation. We describe here a rare case of juvenile granulosa cell tumor in a female with central precocious puberty (CPP. An 8-year old girl with CPP presented with vaginal bleeding four months after the diagnosis and before starting treatment with gonadotropin-releasing hormone (GnRH-analogs. Suppression of basal follicle-stimulating hormone (FSH level, elevation of serum estradiol, progesterone and Cancer Antigen-125 were documented. Abdominal ultrasound examination (US and magnetic resonance imaging showed a pelvic mass affecting the left ovary. A left salpingo-oophorectomy was performed and the mass was totally resected. Juvenile granulosa cell ovarian tumor was diagnosed. One month post surgery, estradiol and progesterone decreased to values of the first evaluation and FSH increased; Cancer Antigen-125 resulted normal while ultrasound pelvic examination showed absence of pelvic masses. In our patient, the tumor had grown very quickly since hormonal data demonstrated a CPP without any evidence of ovarian mass on US only four months before diagnosis. The overstimulation of the FSH or aberrant activation of FSH receptors may have contributed to the development of the mass.

  14. THE EFFECT OF CURCUMIN ON SECRETORY ACTIVITY, PROLIFERATION AND APOPTOSIS OF THE PORCINE OVARIAN GRANULOSA CELLS

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    Attila Kádasi

    2012-08-01

    Full Text Available The aim of this in vitro study was to examine the effect of natural plant (Curcuma longa molecule curcumin on secretory activity, proliferation and apoptosis of porcine granulosa cells. The secretion of steroid hormones (progesterone, testosterone, accumulation of PCNA (marker of proliferation and bax (marker of apoptosis in granulosa cells of swine ovaries after curcumin treatment at the doses 0, 1, 10, 100 μg.mL-1 was determined by RIA and immunocytochemistry. It was observed that, addition of curcumin stimulated progesterone (at doses 1 and 10 μg.mL-1, but not 100 μg.mL-1 and testosterone at (100 μg.mL-1 but not 1 and 10 μg.mL-1 release. The number of cells contained PCNA was down-regulated by curcumin administration (at dose of 10 μg.mL-1, but not of 1 and 100 μg.mL-1. Bax expression was stimulated by curcumin at all doses added. Our results suggest a direct effect of curcumin on ovarian functions: steroidogenesis, proliferation and apoptosis. This could suggest antireproductive properties of curcumin in swine ovaries.

  15. Redundant Roles of SMAD2 and SMAD3 in Ovarian Granulosa Cells In Vivo ▿

    Science.gov (United States)

    Li, Qinglei; Pangas, Stephanie A.; Jorgez, Carolina J.; Graff, Jonathan M.; Weinstein, Michael; Matzuk, Martin M.

    2008-01-01

    Transforming growth factor β (TGF-β) superfamily members are critical in maintaining cell growth and differentiation in the ovary. Although signaling of activins, TGF-βs, growth differentiation factor 9, and nodal converge preferentially to SMAD2 and SMAD3, the in vivo functions and redundancy of these SMADs in the ovary and female reproduction remain largely unidentified. To circumvent the deleterious phenotypic aspects of ubiquitous deletion of Smad2 and Smad3, a conditional knockout strategy was formulated to selectively inactivate Smad2, Smad3, or both Smad2 and Smad3 in ovarian granulosa cells. While granulosa cell ablation of individual Smad2 or Smad3 caused insignificant changes in female fertility, deletion of both Smad2 and Smad3 led to dramatically reduced female fertility and fecundity. These defects were associated with the disruption of multiple ovarian processes, including follicular development, ovulation, and cumulus cell expansion. Furthermore, the impaired expansion of cumulus cells may be partially associated with altered cumulus expansion-related transcripts that are regulated by SMAD2/3 signaling. Our results indicate that SMAD2 and SMAD3 function redundantly in vivo to maintain normal female fertility and further support the involvement of an intraovarian SMAD2/3 pathway in mediating oocyte-produced signals essential for coordinating key events of the ovulatory process. PMID:18809571

  16. Paracrine Regulation of Steroidogenesis in Theca Cells by Granulosa Cells Derived from Mouse Preantral Follicles.

    Science.gov (United States)

    Liu, Xiaoqiang; Qiao, Pengyun; Jiang, Aifang; Jiang, Junyi; Han, Haiyan; Wang, Li; Ren, Chune

    2015-01-01

    Interaction partners of follicular cells play a significant role in steroidogenesis, follicular formation, and development. Androgen secreted by theca cells (TCs) can initiate follicle development and ovulation and provide precursor materials for estrogen synthesis. Therefore, studies on ovarian microenvironment will not only lead to better understanding of the steroidogenesis but also have clinical significance for ovarian endocrine abnormalities such as hyperandrogenism in polycystic ovary syndrome (PCOS). This study applied the Transwell coculture model to investigate if the interaction between granulosa and theca cells may affect androgen production in theca cells. Concentrations of testosterone and androstenedione in the spent medium were measured by radioimmunoassay and enzyme linked immunosorbent assay, respectively. The results show that the coculture with granulosa cells (GCs) increases steroidogenesis in TCs. In addition, testosterone and androstenedione productions in response to LH stimulation were also increased in the coculture model. Significantly increased mRNA expressions of steroidogenic enzymes (Star, Cyp11a1, Cyp17a1, and Hsd3b2) were observed in the cocultured TCs. Thus, GCs were capable of promoting steroidogenesis and LH responsiveness in TCs. This study provided a basis for further exploration of ovarian endocrine mechanism and pathologies.

  17. Paracrine Regulation of Steroidogenesis in Theca Cells by Granulosa Cells Derived from Mouse Preantral Follicles

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    Xiaoqiang Liu

    2015-01-01

    Full Text Available Interaction partners of follicular cells play a significant role in steroidogenesis, follicular formation, and development. Androgen secreted by theca cells (TCs can initiate follicle development and ovulation and provide precursor materials for estrogen synthesis. Therefore, studies on ovarian microenvironment will not only lead to better understanding of the steroidogenesis but also have clinical significance for ovarian endocrine abnormalities such as hyperandrogenism in polycystic ovary syndrome (PCOS. This study applied the Transwell coculture model to investigate if the interaction between granulosa and theca cells may affect androgen production in theca cells. Concentrations of testosterone and androstenedione in the spent medium were measured by radioimmunoassay and enzyme linked immunosorbent assay, respectively. The results show that the coculture with granulosa cells (GCs increases steroidogenesis in TCs. In addition, testosterone and androstenedione productions in response to LH stimulation were also increased in the coculture model. Significantly increased mRNA expressions of steroidogenic enzymes (Star, Cyp11a1, Cyp17a1, and Hsd3b2 were observed in the cocultured TCs. Thus, GCs were capable of promoting steroidogenesis and LH responsiveness in TCs. This study provided a basis for further exploration of ovarian endocrine mechanism and pathologies.

  18. [Secondary amenorrhea and LH hypersecretion. An unusual report of a granulosa cell ovarian tumor].

    Science.gov (United States)

    Arteaga, E; Campusano, C; Fernández, C

    1993-04-01

    Granulosa cell ovarian tumors are infrequent. Since they originate from the gonadal stroma, they retain a high secretory potential and some of their clinical manifestations may be secondary to the production of sexual steroids. A 36 year old woman with an ovarian tumor presenting as a secondary amenorrhea is reported. This patient had a positive progesterone test and her hormonal profile showed a maintained LH hypersecretion (> 75 mUl/ml) which, joined to the presence of a hypophyseal microadenoma lead to suspect the presence of a gonadotrophin secreting tumor. The absence of LH response to TRH and its adequate suppression using oral contraceptives discarded this diagnosis. The histopathology of the excised ovarian tumor demonstrated that it is was a granulosa cell tumor. The physiopathological explanation of the case is based on the maintained levels of estrogens produced by the tumor that, through a positive feed-back mechanism similar to that of the polycystic ovary syndrome, produced a tonic LH elevation and GnRH hyper response. After the tumor excision, ovulatory cycles resumed and the patient became pregnant, facts that confirm the postulated hypothesis.

  19. GGPP-Mediated Protein Geranylgeranylation in Oocyte Is Essential for the Establishment of Oocyte-Granulosa Cell Communication and Primary-Secondary Follicle Transition in Mouse Ovary.

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    Chen Jiang

    2017-01-01

    Full Text Available Folliculogenesis is a progressive and highly regulated process, which is essential to provide ova for later reproductive life, requires the bidirectional communication between the oocyte and granulosa cells. This physical connection-mediated communication conveys not only the signals from the oocyte to granulosa cells that regulate their proliferation but also metabolites from the granulosa cells to the oocyte for biosynthesis. However, the underlying mechanism of establishing this communication is largely unknown. Here, we report that oocyte geranylgeranyl diphosphate (GGPP, a metabolic intermediate involved in protein geranylgeranylation, is required to establish the oocyte-granulosa cell communication. GGPP and geranylgeranyl diphosphate synthase (Ggpps levels in oocytes increased during early follicular development. The selective depletion of GGPP in mouse oocytes impaired the proliferation of granulosa cells, primary-secondary follicle transition and female fertility. Mechanistically, GGPP depletion inhibited Rho GTPase geranylgeranylation and its GTPase activity, which was responsible for the accumulation of cell junction proteins in the oocyte cytoplasm and the failure to maintain physical connection between oocyte and granulosa cells. GGPP ablation also blocked Rab27a geranylgeranylation, which might account for the impaired secretion of oocyte materials such as Gdf9. Moreover, GGPP administration restored the defects in oocyte-granulosa cell contact, granulosa cell proliferation and primary-secondary follicle transition in Ggpps depletion mice. Our study provides the evidence that GGPP-mediated protein geranylgeranylation contributes to the establishment of oocyte-granulosa cell communication and then regulates the primary-secondary follicle transition, a key phase of folliculogenesis essential for female reproductive function.

  20. A rare case of ovarian granulosa cell tumour in an adolescent girl with secondary amenorrhea and virilisation

    Directory of Open Access Journals (Sweden)

    Arun Nayak

    2015-10-01

    Full Text Available Sex cord stromal ovarian tumors account for about 5-8% of all ovarian malignancies. Granulosa cell tumor is an estrogen secreting low grade malignant tumor and is seen in women of all ages with 2% bilaterality. Based upon histology, these neoplasms can be classified as adult and juvenile Granulosa cell tumors. Pre-pubertal lesions present clinically with sexual pseudo precocity whereas post-pubertal lesions present with menstrual irregularities or secondary amenorrhea or postmenopausal bleeding. This case report presents a rare case of Granulosa cell tumor with clinical presentation mimicking virilizing ovarian neoplasm in a 19 year old girl with history of abdominal swelling, secondary amenorrhea and virilization since one year duration. Clinical examination revealed a 20-22 weeks size abdominal mass, cystic in consistency, arising from pelvis and CT scan suggested presence of mucinous cystadenoma with solid areas with suspected malignancy. Exploratory laparotomy with right salpingo-oophorectomy was performed with staging as tumor was found confined to ovary with no involvement of other ovary or any other pelvic organ or peritoneum. Therefore, disease was staged as stage 1A. Histopathological examination confirmed the presence of Granulosa cell tumor with fibrothecomatous areas. [Int J Reprod Contracept Obstet Gynecol 2015; 4(5.000: 1653-1656

  1. Knockdown of XBP1 by RNAi in Mouse Granulosa Cells Promotes Apoptosis, Inhibits Cell Cycle, and Decreases Estradiol Synthesis

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    Nan Wang

    2017-05-01

    Full Text Available Granulosa cells are crucial for follicular growth, development, and follicular atresia. X-box binding protein 1 (XBP1, a basic region-leucine zipper protein, is widely involved in cell differentiation, proliferation, apoptosis, cellular stress response, and other signaling pathways. In this study, RNA interference, flow cytometry, western blot, real-time PCR, Cell Counting Kit (CCK8, and ELISA were used to investigate the effect of XBP1 on steroidogenesis, apoptosis, cell cycle, and proliferation of mouse granulosa cells. ELISA analysis showed that XBP1 depletion significantly decreased the concentrations of estradiol (E2. Additionally, the expression of estrogen synthesis enzyme Cyp19a1 was sharply downregulated. Moreover, flow cytometry showed that knockdown of XBP1 increased the apoptosis rate and arrests the cell cycle in S-phase in granulosa cells (GCs. Further study confirmed these results. The expression of CCAAT-enhancer-binding protein homologous protein (CHOP, cysteinyl aspartate specific proteases-3 (caspase-3, cleaved caspase-3, and Cyclin E was upregulated, while that of Bcl-2, Cyclin A1, and Cyclin B1 was downregulated. Simultaneously, CCK8 analysis indicated that XBP1 disruption inhibited cell proliferation. In addition, XBP1 knockdown also alters the expression of Has2 and Ptgs2, two essential genes for folliculogenesis. Collectively, these data reveal a novel critical role of XBP1 in folliculogenesis by regulating the cell cycle, apoptosis, and steroid synthesis of mouse granulosa cells.

  2. Evidence for Existence of Immunoglobulins that Block Ovarian Granulosa Cell Growth in Vitro. A Putative Role in Resistant Ovary Syndrome?

    NARCIS (Netherlands)

    WEISSENBRUCH, MIRJAM M. van; HOEK, ANNEMIEKE; VLIET-BLEEKER, INGRID van; SCHOEMAKER, JOOP; DREXHAGE, HEMMO

    1991-01-01

    The sera of 26 patients with premature ovarian failure were examined in order to detect immunoglobulin-G (IgGs) that can block FSH-induced in vitro granulosa cell DNA synthesis via, a Feulgen cytochemical bioassay system. The IgGs of four patients with polycystic ovary-like disease, five postmenopau

  3. Expression Levels of PPARγ and CYP-19 in Polycystic Ovarian Syndrome Primary Granulosa Cells: Influence of ω-3 Fatty Acid

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    Mina Zaree

    2015-07-01

    Full Text Available Background: The omega-3 fatty acid (ω-3 fatty acid such as eicosapentaenoic acid (EPA is currently used in the clinic as a nutritional supplement in the treatment of polycystic ovarian syndrome (PCOS. The present study was designed to investigate the effect of EPA on the expression levels of peroxisome proliferator-activated receptor gamma (PPARγ and cytochrome P450 aromatase (encoded by the CYP-19 in primary cultured granulosa cells (GC from patients undergoing in vitro fertilization (IVF, and also to compare these effects with those in GC of PCOS patients. Materials and Methods: In this experimental study, human GC were isolated, primary cultured in vitro, exposed to a range of concentrations of the EPA and investigated with respect to gene expression levels of PPARγ and CYP-19 using real time-polymerase chain reaction (PCR. The participants (n=30 were the patients admitted to the IVF Center in February-March 2013 at Alzahra Hospital, Tabriz, Iran, who were divided into two groups as PCOS (n=15 and non-PCOS (n=15 women (controls. Results: All doses of the EPA significantly induced PPARγ mRNA gene expression level as compared to the control recombinant follicle stimulating hormone (rFSH alone condition. High doses of EPA in the presence of rFSH produced a stimulatory effect on expression level of PPARγ (2.15-fold, P=0.001 and a suppressive effect (0.56-fold, P=0.01 on the expression level of CYP-19, only in the PCOS GC. Conclusion: EPA and FSH signaling pathway affect differentially on the gene expression levels of PPARγ and CYP-19 in PCOS GC. Altered FSH-induced PPARγ activity in PCOS GC may modulate the CYP-19 gene expression in response to EPA, and possibly modulates the subsequent steroidogenesis of these cells.

  4. Structural and spectroscopic features of lutein/butanoyl-β-cyclodextrin nanoassemblies

    DEFF Research Database (Denmark)

    Stancanelli, R.; Løjkner, L.D.; Larsen, Kim Lambertsen

    2012-01-01

    Lutein, the primary carotenoid present in the central area of the retina of eye appears to be associated with the protection against age-related macular degeneration (the leading cause of blindness in older adults). Its lipophilicity and consequently its scarce water solubility (1.3 × 10−9 M) rep......–vis measurements revealed that electronic properties of lutein were maintained when interact with ACyD nanoaggregates. The monitoring of the entapped carotenoid leaking from ACyD nanostructures was investigated suggesting the potential of Lut/ACyD nanoassemblies in drug delivery....

  5. Tumores das células da granulosa dos ovários: estudo de 24 casos Granulosa cell tumors of the ovary: a study of 24 cases

    Directory of Open Access Journals (Sweden)

    Artur Lício Rocha Bezerra

    2004-09-01

    Full Text Available OBJETIVOS: avaliar estadiamento, procedimentos cirúrgicos e evolução dos tumores das células da granulosa (TCG dos ovários, dando ênfase à possibilidade de cirurgia conservadora (ooforectomia unilateral. MÉTODOS: este é um estudo de coorte retrospectivo no qual foram incluídas 24 pacientes com TCG tratadas no período de janeiro de 1994 a janeiro de 2004. Foram analisadas variáveis de interesse, tais como idade da paciente, sintomatologia, tamanho tumoral ao exame físico, estadiamento, modalidades de tratamento (tipos de cirurgia e adjuvância com quimioterapia e/ou radioterapia e prognóstico. As associações entre as variáveis foram avaliadas pelo teste do chi-quadrado e para significância estatística considerou-se p OBJECTIVE: to evaluate staging, surgical procedures and outcome of ovarian granulosa cell tumors (GCT, with emphasis in the possibility of conservative surgery (unilateral ooforectomy. METHODS: this is a retrospective cohort study in which 24 patients treated with GCT during the period of January 1994 to January 2004 were included. Variables of interest such as patient's age, symptoms, tumor size on physical examination, staging, treatment modalities (types of surgery and of adjuvant chemotherapy and/or radiotherapy and prognosis have been analyzed. The chi-square test was used to check the association between variables, and the level of significance was set at p < 0.05, always describing the two-tailed Fisher exact test. RESULTS: the patients' age varied from 30 to 82 (mean 51.7 years old. The most frequently referred symptom was pelvic pain (n = 10; 41.7%. Fourteen patients (58.3% presented clinical stage (CS Ic, 5 (20.8% CS IIIc and 5 (20.8% CS Ia. More advanced clinical stages were significantly associated with palpable pelvic masses (p < 0.04. Endometrial hyperplasias were found in 3 (25% of the 12 hysterectomy specimens. The follow-up period varied from 2 to 114 (mean 42.5 months. Among the 16 (66

  6. Regulatory role of kit ligand-c-kit interaction and oocyte factors in steroidogenesis by rat granulosa cells.

    Science.gov (United States)

    Miyoshi, Tomoko; Otsuka, Fumio; Nakamura, Eri; Inagaki, Kenichi; Ogura-Ochi, Kanako; Tsukamoto, Naoko; Takeda, Masaya; Makino, Hirofumi

    2012-07-06

    Although kit ligand (KL)-c-kit interaction is known to be critical for oogenesis and folliculogenesis, its role in ovarian steroidogenesis has yet to be elucidated. We studied the impact of KL-c-kit interaction in regulation of steroidogenesis using rat oocyte/granulosa cell co-culture. In the presence of oocytes, soluble KL suppressed FSH-induced estradiol production and aromatase mRNA expression without affecting FSH-induced progesterone production. The KL effect on steroidogenesis was interrupted by an anti-c-kit neutralizing antibody, suggesting that KL-c-kit interaction is involved in suppression of estrogen by granulosa cells through oocyte c-kit action. The cAMP-PKA pathway activity was not directly involved in the estrogen regulation by KL-c-kit action. It was of note that KL treatment increased the expression levels of oocyte-derived FGF-8, GDF-9 and BMP-6, while it reduced the expression levels of oocyte-derived BMP-15 in the oocyte-granulosa cell co-culture. Given the findings that FGF-8, but not GDF-9, BMP-6 or -15, suppressed FSH-induced estrogen production by granulosa cells, oocyte-derived FGF-8 is linked to suppression of FSH-induced estrogen production through the KL-c-kit interaction. Furthermore, the suppression of FSH-induced estrogen production by KL in the co-culture was reversed by a FGF receptor kinase inhibitor and the effect of the inhibitor was enhanced in combination with extracellular-domain protein of BMPRII, which interferes with BMP-15 and GDF-9 activities. Thus, the actions of endogenous oocyte factors including FGF-8 and BMP-15/GDF-9 were involved in the KL activity that inhibited FSH-induced estradiol production. Collectively, the results indicate that KL-c-kit interaction plays a role in estrogenic regulation through oocyte-granulosa cell communication.

  7. Presence of encircling granulosa cells protects against oxidative stress-induced apoptosis in rat eggs cultured in vitro.

    Science.gov (United States)

    Tiwari, Meenakshi; Tripathi, Anima; Chaube, Shail K

    2017-01-01

    Increased oxidative stress (OS) due to in vitro culture conditions can affect the quality of denuded eggs during various assisted reproductive technologies (ARTs). Presence of intact granulosa cells may protect eggs from OS damage under in vitro culture conditions. The present study was aimed to investigate whether encircling granulosa cells could protect against hydrogen peroxide (H2O2)-induced egg apoptosis in ovulated cumulus oocyte complexes (COCs) cultured in vitro. The OS was induced by exposing COCs as well as denuded eggs with various concentrations of H2O2 for 3 h in vitro. The morphological changes, total reactive oxygen species (ROS) as well as catalase expression, Bax/Bcl-2, cytochrome c levels and DNA fragmentation were analysed in COCs as well as denuded eggs. Our results suggest that H2O2 treatment induced morphological apoptotic features in a concentration-dependent manner in denuded eggs cultured in vitro. The 20 µM of H2O2 treatment induced OS by elevating total ROS level, reduced catalase and Bcl-2 expression levels with overexpression of Bax and cytochrome c and induced DNA fragmentation in denuded eggs cultured in vitro. The presence of encircling granulosa cells protected H2O2-induced morphological apoptotic features by preventing the increase of Bax, cytochrome c expression levels and DNA fragmentation in associated egg. However, 20 µM of H2O2 was sufficient to induce peripheral granulosa cell apoptosis in COCs and degeneration in few denuded eggs cultured in vitro. Taken together our data suggest that the presence of encircling granulosa cells could be beneficial to protect ovulated eggs from OS damage under in vitro culture conditions during various ART programs.

  8. Interleukin-1 (IL-1 system gene expression in granulosa cells: kinetics during terminal preovulatory follicle maturation in the mare

    Directory of Open Access Journals (Sweden)

    Gérard Nadine

    2003-05-01

    Full Text Available Abstract Background A growing body of evidences suggests that the ovary is a site of inflammatory reactions, and thus, ovarian cells could represent sources and targets of the interleukin-1 (IL-1 system. The purpose of this study was to examine the IL-1 system gene expressions in equine granulosa cells, and to study the IL-1β content in follicular fluid during the follicle maturation. For this purpose, granulosa cells and follicular fluids were collected from the largest follicle at the early dominance stage (diameter 24 ± 3 mm or during the preovulatory maturation phase, at T0 h, T6 h, T12 h, T24 h and T34 h after induction of ovulation. Cells were analysed by RT-PCR and follicular fluids were studied by gel electrophoresis and immunoblotting. Results We demonstrated that interleukin-1β (IL-1β, interleukin-1 receptor 2 (IL-1R2 and interleukin-1 receptor antagonist (IL-1RA genes are expressed in equine granulosa cells. We observed that the IL-1β and IL-1RA mRNA content changed in granulosa cells during the terminal follicular maturation whereas IL-1R2 mRNA did not vary. In follicular fluid, IL-1β content fluctuated few hours after induction of ovulation. Conclusions The expression of IL-1β gene in granulosa cells and the follicular fluid IL-1β content seem to be regulated by gonadotropins suggesting that IL-1β could be an intermediate paracrine factor involved in ovulation.

  9. Radioimmunoassay of serum follicle-stimulating hormone and luteinizing hormone in the bottlenosed dolphin.

    Science.gov (United States)

    Schneyer, A; Castro, A; Odell, D

    1985-11-01

    Commercially available radioimmunoassay (RIA) kits for human follicle-stimulating hormone (FSH) and luteinizing hormone (LH) were adapted for quantitation of these hormones in serum from bottlenosed dolphins (Tursiops truncatus). Serum samples from over 160 wild and 70 captive animals were assayed in order to determine basal concentrations of FSH and LH in these animals, as well as to detect possible differences between various groups. Mean FSH and LH levels for all animals were 0.22 +/- 0.08 and 0.37 +/- 0.18 ng/ml, respectively. Although wild animals had higher FSH and LH levels than captive ones, the differences were not statistically significant (P less than 0.07). However, both FSH and LH were significantly (P less than 0.01 and P less than 0.05, respectively) elevated in females when compared to males. Adults and peripubescent animals had significantly (P less than 0.01) higher LH levels than did juveniles. Among wild animals, serum concentrations of FSH and LH reflected seasonal differences. Samples obtained in early summer (Gulf of Mexico population) contained significantly (P less than 0.01) higher concentrations of FSH and LH than samples obtained in the fall (Indian River, Florida population). Both FSH and LH were significantly elevated in samples from confirmed pregnant animals as compared to the overall mean and to a sample from a confirmed nonpregnant female. Our observations indicate that these RIAs can reliably detect serum FSH and LH from bottlenosed dolphins and represent the first quantitation of these hormones in cetaceans.

  10. Purinergic receptor-mediated intracellular Ca2+ oscillations in chicken granulosa cells.

    Science.gov (United States)

    Morley, P; Vanderhyden, B C; Tremblay, R; Mealing, G A; Durkin, J P; Whitfield, J F

    1994-03-01

    These studies were designed to investigate the effects of extracellular ATP on intracellular calcium ion concentration ([Ca2+]i) and progesterone secretion in granulosa cells obtained from the two largest preovulatory follicles (F1 and F2) of hens. [Ca2+]i was measured in cells loaded with the Ca(2+)-responsive fluorescent dye fura-2. The resting [Ca2+]i in these cells was 99 +/- 7 nM (n = 22). There was a 5.7 +/- 0.7-fold increase in [Ca2+]i in all (n = 140) of the cells within 5 sec of adding a maximally stimulatory concentration (100 microM) of extracellular ATP. The initial spike was followed by [Ca2+]i oscillations that returned to the resting level between spikes. The frequency and amplitude of the [Ca2+]i oscillations were varied and persisted for 1-40 min. [Ca2+]i oscillations were also triggered by 100 microM UTP, UDP, GTP, GDP, ADP, and the nonhydrolyzable analog ATP gamma S. Adenosine, AMP, GMP, and UMP (all at 100 microM) were ineffective. The lowest ATP concentration to trigger a [Ca2+]i response was 1 microM. The sustained oscillatory phase of the response, but not the initial spike, was inhibited by incubating the cells in Ca(2+)-free medium containing 2 mM EGTA. The nucleotide-triggered [Ca2+]i oscillations were not affected by adding the dihydropyridine Ca2+ channel blockers verapamil (100 microM), methoxy-verapamil (D600; 100 microM), or nifedipine (10 microM), before or during the response. However, the oscillations, but not the initial spike, were prevented by pretreating the cells with a general Ca2+ channel blocker, lanthanum (1 mM) or cobalt (5 mM). Lanthanum and cobalt also promptly stopped the [Ca2+]i oscillations when added during the oscillatory phase. The nucleotide-triggered [Ca2+]i response was also abolished by pretreating the cells with an inhibitor of inositol phospholipid hydrolysis, neomycin (1.5 mM). In 3-h incubations, adenosine (100 microM) or ATP (100 microM) did not affect basal or LH (20 or 100 ng/ml)-stimulated progesterone

  11. Structural and spectroscopic features of lutein/butanoyl-β-cyclodextrin nanoassemblies.

    Science.gov (United States)

    Stancanelli, Rosanna; Løjkner, Lars Damgaard; Larsen, Kim Lambertsen; Guardo, Marta; Cannavà, Carmela; Tommasini, Silvana; Ventura, Cinzia Anna; Calabrò, Maria Luisa; Micali, Norberto; Villari, Valentina; Mazzaglia, Antonino

    2012-12-01

    Lutein, the primary carotenoid present in the central area of the retina of eye appears to be associated with the protection against age-related macular degeneration (the leading cause of blindness in older adults). Its lipophilicity and consequently its scarce water solubility (1.3×10(-9)M) represent a drawback for bioavailability. To circumvent these unfavorable characteristics, in this work lutein (Lut) have been encapsulated in amphiphilic cyclodextrin (ACyD) by following the well-established strategy of entrapping a lipophilic drug in CyD carriers. Primary face butyrate modified β-cyclodextrins (C(4:7)) form in water nanoaggregates with a average size of 250nm and a ζ-potential of about -6mV. They are able to entrap lutein at 1:6 Lut/ACyD molar ratio by yielding nanoassemblies of vesicular aspect (320nm and -8mV) such as observed by static, dynamic and electrophoretic light-scattering. UV-vis measurements revealed that electronic properties of lutein were maintained when interact with ACyD nanoaggregates. The monitoring of the entapped carotenoid leaking from ACyD nanostructures was investigated suggesting the potential of Lut/ACyD nanoassemblies in drug delivery.

  12. Clearance of yellow pigments lutein and zeathanxin in channel catfish reared at different water temperatures

    Science.gov (United States)

    A study was conducted to determine clearance time of yellow pigments lutein and zeaxanthin in channel catfish at various temperatures. Fish of initial weight of 13.4 g were stocked into flow-through aquaria and fed once daily with a yellow pigment enhanced diet for 11 weeks when the yellow color be...

  13. Luteinizing hormone-releasing hormone receptor antagonist may reduce postmenopausal flushing

    NARCIS (Netherlands)

    Gastel, P. van; Zanden, M. van der; Telting, D.; Filius, M.; Bancsi, L.; Boer, H. de

    2012-01-01

    OBJECTIVE: Hormone therapy (HT) is the most effective treatment of postmenopausal (PMP) flushing; however, its use is often contraindicated. As an alternative option, we explored the efficacy of the luteinizing hormone-releasing hormone (LHRH) receptor antagonist cetrorelix in women with severe PMP

  14. Enhanced Productivity of a Lutein-Enriched Novel Acidophile Microalga Grown on Urea

    NARCIS (Netherlands)

    Casal, C.; Cuaresma, M.; Vega, J.M.; Vilchez, C.

    2011-01-01

    Coccomyxa acidophila is an extremophile eukaryotic microalga isolated from the Tinto River mining area in Huelva, Spain. Coccomyxa acidophila accumulates relevant amounts of b-carotene and lutein, well-known carotenoids with many biotechnological applications, especially in food and health-related

  15. Two stage heterotrophy/photoinduction culture of Scenedesmus incrassatulus: potential for lutein production.

    Science.gov (United States)

    Flórez-Miranda, Liliana; Cañizares-Villanueva, Rosa Olivia; Melchy-Antonio, Orlando; Jerónimo, Fernando Martínez-; Flores-Ortíz, Cesar Mateo

    2017-09-16

    A biomass production process including two stages, heterotrophy/photoinduction (TSHP), was developed to improve biomass and lutein production by the green microalgae Scenedesmus incrassatulus. To determine the effects of different nitrogen sources (yeast extract and urea) and temperature in the heterotrophic stage, experiments using shake flask cultures with glucose as the carbon source were carried out. The highest biomass productivity and specific pigment concentrations were reached using urea+vitamins (U+V) at 30°C. The first stage of the TSHP process was done in a 6L bioreactor, and the inductions in a 3L airlift photobioreactor. At the end of the heterotrophic stage, S. incrassatulus achieved the maximal biomass concentration, increasing from 7.22gL(-1) to 17.98gL(-1) with an increase in initial glucose concentration from 10.6gL(-1) to 30.3gL(-1). However, the higher initial glucose concentration resulted in a lower specific growth rate (μ) and lower cell yield (Yx/s), possibly due to substrate inhibition. After 24h of photoinduction, lutein content in S. incrassatulus biomass was 7 times higher than that obtained at the end of heterotrophic cultivation, and the lutein productivity was 1.6 times higher compared with autotrophic culture of this microalga. Hence, the two-stage heterotrophy/photoinduction culture is an effective strategy for high cell density and lutein production in S. incrassatulus. Copyright © 2017. Published by Elsevier B.V.

  16. Enhanced Productivity of a Lutein-Enriched Novel Acidophile Microalga Grown on Urea

    NARCIS (Netherlands)

    Casal, C.; Cuaresma, M.; Vega, J.M.; Vilchez, C.

    2011-01-01

    Coccomyxa acidophila is an extremophile eukaryotic microalga isolated from the Tinto River mining area in Huelva, Spain. Coccomyxa acidophila accumulates relevant amounts of b-carotene and lutein, well-known carotenoids with many biotechnological applications, especially in food and health-related i

  17. Development of the membrana granulosa of bovine antral follicles: structure, location of mitosis and pyknosis, and immunolocalization of involucrin and vimentin.

    Science.gov (United States)

    van Wezel, I L; Krupa, M; Rodgers, R J

    1999-01-01

    The membrana granulosa of the ovarian follicle is termed the 'follicular epithelium', yet there have been no studies considering its epithelial nature and how it changes during follicular development. Therefore, these issues were investigated using histology (n = 45 ovaries), considering its structure and the location of proliferating and dying cells, and drawing analogies with other epithelia. Additionally, differences between the layers of granulosa cells were demonstrated by immunohistochemistry (n = 7 ovaries). The structure of the membrana granulosa differed between follicles. Six arbitrary classifications were designed based on these structures, 80 follicles were allocated (n = 13 ovaries) to these classes and the follicular diameters were then measured. For the first time, differences in membrana granulosa structure were shown to correspond to follicle size. Follicles in classes 1-3, where basal granulosa cells were columnar with nuclei positioned basally in the cell, were all 5 mm had only rounded basal cells. In all these classes, cells in the middle zone were rounded; cells aligning the antrum were often flattened. Irrespective of follicle class, cell proliferation and cell death were shown to be predominantly in the middle portions, rather than the most antral or most basal portions, of the membrana granulosa of healthy and atretic follicles. Involucrin, a marker of keratinocyte differentiation, was localized to the suprabasal region of the membrana granulosa of healthy follicles, particularly in the second and third cellular layers in from the follicular basal lamina. Conversely, the staining intensity for the intermediate filament protein vimentin was lowest in this region, and greatest in the more antral and basal regions. In atretic follicles, there was widespread staining for involucrin and vimentin throughout the membrana granulosa. In conclusion, the membrana granulosa is highly structured, and alters with follicular development. Layers in the

  18. MicroRNA-183-96-182 Cluster Regulates Bovine Granulosa Cell Proliferation and Cell Cycle Transition by Coordinately Targeting FOXO1.

    Science.gov (United States)

    Gebremedhn, Samuel; Salilew-Wondim, Dessie; Hoelker, Michael; Rings, Franca; Neuhoff, Christiane; Tholen, Ernst; Schellander, Karl; Tesfaye, Dawit

    2016-06-01

    Large-scale expression profiling of micro-RNAs (miRNAs) in bovine granulosa cells from dominant and subordinate follicles on Day 19 of the estrous cycle revealed enriched micro-RNA-183-96-182 cluster miRNAs in preovulatory dominant follicles that coordinately regulate the forkhead box protein O1 (FOXO1) gene. However, little is known about the role of this cluster in bovine granulosa cell function. We used an in vitro granulosa cell culture model to investigate this role. Granulosa cells aspirated from small growing follicles (3-5 mm in diameter) were cultured in Dulbecco modified Eagle medium/F-12 medium supplemented with fetal bovine serum and transfected with locked nucleic acid-based miRNA mimics, inhibitors, and corresponding negative controls. Overexpression of the miRNA cluster resulted in suppression of FOXO1 mRNA and protein, whereas inhibition of the cluster increased expression of FOXO1 mRNA. Overexpression also increased the relative rate of cell proliferation, whereas inhibition slowed it down. Similarly, the proportion of cells under G0/G1 arrest declined, whereas the ratio of cells in S phase increased in response to miR-183-96-182 overexpression. Selective knockdown of FOXO1 mRNA using anti-FOXO1 small interfering RNA increased the rate of granulosa cell proliferation, decreased the proportion of cells under G0/G1 arrest, and increased the proportion of cells in the S phase of cell cycle. Our data suggest that miR-183-96-182 cluster miRNAs promote proliferation and G1/S transition of bovine granulosa cells by coordinately targeting FOXO1, suggesting a critical role in granulosa cell function. MicroRNA-183-96-182 cluster regulates bovine granulosa cell function by targeting FOXO1 gene.

  19. Positive effect of dietary lutein and cholesterol on the undirected song activity of an opportunistic breeder

    Directory of Open Access Journals (Sweden)

    Stefania Casagrande

    2016-10-01

    Full Text Available Song is a sexually selected trait that is thought to be an honest signal of the health condition of an individual in many bird species. For species that breed opportunistically, the quantity of food may be a determinant of singing activity. However, it is not yet known whether the quality of food plays an important role in this respect. The aim of the present study was to experimentally investigate the role of two calorie-free nutrients (lutein and cholesterol in determining the expression of a sexually selected behavior (song rate and other behaviors (locomotor activity, self-maintenance activity, eating and resting in male zebra finches (Taeniopygia guttata. We predicted that males supplemented with lutein and cholesterol would sing at higher rates than controls because both lutein and cholesterol have important health-related physiological functions in birds and birdsong mirrors individual condition. To control for testosterone secretion that may upregulate birdsong, birds were exposed to a decreasing photoperiod. Our results showed that control males down-regulated testosterone in response to a decreasing photoperiod, while birds treated with lutein or cholesterol maintained a constant singing activity. Both lutein- and cholesterol-supplemented groups sang more than control groups by the end of the experiment, indicating that the quality of food can affect undirected song irrespective of circulating testosterone concentrations. None of the other measured behaviors were affected by the treatment, suggesting that, when individuals have full availability of food, sexually selected song traits are more sensitive to the effect of food quality than other behavioral traits. Overall the results support our prediction that undirected song produced by male zebra finches signals access to high-quality food.

  20. Positive effect of dietary lutein and cholesterol on the undirected song activity of an opportunistic breeder

    Science.gov (United States)

    Pinxten, Rianne; Zaid, Erika; Eens, Marcel

    2016-01-01

    Song is a sexually selected trait that is thought to be an honest signal of the health condition of an individual in many bird species. For species that breed opportunistically, the quantity of food may be a determinant of singing activity. However, it is not yet known whether the quality of food plays an important role in this respect. The aim of the present study was to experimentally investigate the role of two calorie-free nutrients (lutein and cholesterol) in determining the expression of a sexually selected behavior (song rate) and other behaviors (locomotor activity, self-maintenance activity, eating and resting) in male zebra finches (Taeniopygia guttata). We predicted that males supplemented with lutein and cholesterol would sing at higher rates than controls because both lutein and cholesterol have important health-related physiological functions in birds and birdsong mirrors individual condition. To control for testosterone secretion that may upregulate birdsong, birds were exposed to a decreasing photoperiod. Our results showed that control males down-regulated testosterone in response to a decreasing photoperiod, while birds treated with lutein or cholesterol maintained a constant singing activity. Both lutein- and cholesterol-supplemented groups sang more than control groups by the end of the experiment, indicating that the quality of food can affect undirected song irrespective of circulating testosterone concentrations. None of the other measured behaviors were affected by the treatment, suggesting that, when individuals have full availability of food, sexually selected song traits are more sensitive to the effect of food quality than other behavioral traits. Overall the results support our prediction that undirected song produced by male zebra finches signals access to high-quality food.

  1. Tetrodotoxin levels in larval and metamorphosed newts (Taricha granulosa) and palatability to predatory dragonflies.

    Science.gov (United States)

    Gall, Brian G; Stokes, Amber N; French, Susannah S; Schlepphorst, Elizabeth A; Brodie, Edmund D; Brodie, Edmund D

    2011-06-01

    Some populations of the newt Taricha granulosa possess extremely high concentrations of the neurotoxin tetrodotoxin (TTX). Tetrodotoxin is present in adult newts and their eggs, but has been assumed to be absent from the larval stage. We tested larval and metamorphosed juveniles for the presence of TTX and evaluated the palatability of these developmental stages to predatory dragonfly nymphs. All developmental stages retained substantial quantities of TTX and almost all individuals were unpalatable to dragonfly nymphs. Tetrodotoxin quantity varied greatly among individuals. When adjusted for mass, TTX concentrations declined steadily through metamorphosis. Several juveniles were palatable to dragonflies and these individuals had significantly lower TTX levels than unpalatable juveniles. These results suggest that despite previous assumptions, substantial quantities of TTX, originally deposited in the embryo, are retained by the developing larvae and metamorphosed juveniles and this quantity is enough to make them unpalatable to some potential predators.

  2. Adding of ascorbic acid to the culture medium influences the antioxidant status and some biochemical parameters in the hen granulosa cells.

    Science.gov (United States)

    Capcarova, M; Kolesarova, A; Kalafova, A; Bulla, J; Sirotkin, A V

    2015-07-01

    The aim of the present study was to determine the activity of superoxide dismutase (SOD), total antioxidant status (TAS) of the hen granulosa cells, and selected biochemical parameters, including calcium, phosphorus, sodium, potassium, glucose, cholesterol, proteins, in the culture medium of granulosa cells after exposing them to ascorbic acid in vitro conditions. Ovarian granulosa cells of hens were incubated with various doses of ascorbic acid (E1 0.09 mg/ml, E2 0.13 mg/ml, E3 0.17 mg/ml, E4 0.33 mg/ml, E5 0.5 mg/ml). Ascorbic acid did not manifest antioxidant potential and higher doses of ascorbic acid (0.17; 0.33 and 0.5 mg/ml) decreased the activity of SOD in granulosa cells. Vitamin application resulted in a significantly (p<0.05) higher accumulation of Na+ and K+ in culture media of granulosa cells and decreased the concentration of glucose and proteins. These results indicate that ascorbic acid might be involved in the regulation of selected biochemical and physiological processes in ovarian granulosa cells.

  3. Critical Role of FoxO1 in Granulosa Cell Apoptosis Caused by Oxidative Stress and Protective Effects of Grape Seed Procyanidin B2

    Science.gov (United States)

    Zhang, Jia-Qing; Gao, Bin-Wen; Wang, Jing; Ren, Qiao-Ling; Chen, Jun-Feng; Ma, Qiang; Zhang, Zi-Jing; Xing, Bao-Song

    2016-01-01

    Reactive oxygen species (ROS) are closely related to the follicular granulosa cell apoptosis. Grape seed procyanidin B2 (GSPB2) has been reported to possess potent antioxidant activity. However, the GSPB2-mediated protective effects and the underlying molecular mechanisms in granulosa cell apoptosis process remain unknown. In this study, we showed for the first time that GSPB2 treatment decreased FoxO1 protein level, improved granulosa cell viability, upregulated LC3-II protein level, and reduced granulosa cell apoptosis rate. Under a condition of oxidative stress, GSPB2 reversed FoxO1 nuclear localization and increased its level in cytoplasm. In addition, FoxO1 knockdown inhibited the protective effects of GSPB2 induced. Our findings suggest that FoxO1 plays a pivotal role in regulating autophagy in granulosa cells, GSPB2 exerts a potent and beneficial role in reducing granulosa cell apoptosis and inducing autophagy process, and targeting FoxO1 could be significant in fighting against oxidative stress-reduced female reproductive system diseases. PMID:27057282

  4. Critical Role of FoxO1 in Granulosa Cell Apoptosis Caused by Oxidative Stress and Protective Effects of Grape Seed Procyanidin B2

    Directory of Open Access Journals (Sweden)

    Jia-Qing Zhang

    2016-01-01

    Full Text Available Reactive oxygen species (ROS are closely related to the follicular granulosa cell apoptosis. Grape seed procyanidin B2 (GSPB2 has been reported to possess potent antioxidant activity. However, the GSPB2-mediated protective effects and the underlying molecular mechanisms in granulosa cell apoptosis process remain unknown. In this study, we showed for the first time that GSPB2 treatment decreased FoxO1 protein level, improved granulosa cell viability, upregulated LC3-II protein level, and reduced granulosa cell apoptosis rate. Under a condition of oxidative stress, GSPB2 reversed FoxO1 nuclear localization and increased its level in cytoplasm. In addition, FoxO1 knockdown inhibited the protective effects of GSPB2 induced. Our findings suggest that FoxO1 plays a pivotal role in regulating autophagy in granulosa cells, GSPB2 exerts a potent and beneficial role in reducing granulosa cell apoptosis and inducing autophagy process, and targeting FoxO1 could be significant in fighting against oxidative stress-reduced female reproductive system diseases.

  5. Conditional Deletion of the Retinoblastoma (Rb) Gene in Ovarian Granulosa Cells Leads to Premature Ovarian Failure

    Science.gov (United States)

    Andreu-Vieyra, Claudia; Chen, Ruihong; Matzuk, Martin M.

    2008-01-01

    The retinoblastoma protein (RB) regulates cell proliferation and survival by binding to the E2F family of transcription factors. Recent studies suggest that RB also regulates differentiation in a variety of cell types, including myocytes, neurons, adipocytes, and chondrocytes. Rb mutations have been found in ovarian cancer; however, the role of RB in normal and abnormal ovarian function remains unclear. To test the hypothesis that loss of Rb induces ovarian tumorigenesis, we generated an ovarian granulosa cell conditional knockout of Rb (Rb cKO) using the Cre/lox recombination system. Rb cKO females showed 100% survival and no ovarian tumor formation through 9 months of age, but they developed progressive infertility. Prepubertal Rb cKO females showed increased ovulation rates compared with controls, correlating with increased follicle recruitment, higher Fshr and Kitl mRNA levels, and lower anti-Müllerian hormone levels. In contrast, the ovulation rate of 6-wk-old females was similar to that of controls. Morphometric analysis of Rb cKO ovaries from 6-wk-old and older females showed increased follicular atresia and apoptosis. Rb cKO ovaries and preantral follicles had abnormal levels of known direct and indirect target genes of RB, including Rbl2/p130, E2f1, Ccne2, Myc, Fos, and Tgfb2. In addition, preantral follicles showed increased expression of the granulosa cell differentiation marker Inha, decreased levels of Foxl2 and Cyp19a1 aromatase, and abnormal expression of the nuclear receptors Nr5a1, Nr5a2, and Nr0b1. Taken together, our results suggest that RB is required for the temporal-specific pattern of expression of key genes involved in follicular development. PMID:18599617

  6. Evaluation of steroidogenic capacity after follicle stimulating hormone stimulation in bovine granulosa cells of Revalor 200® implanted heifers

    Institute of Scientific and Technical Information of China (English)

    Andrea DStapp; Craig AGifford; Dennis MHallford; Jennifer AHernandez Gifford

    2014-01-01

    Background:Heifers not used as breeding stock are often implanted with steroids to increase growth efficiency thereby altering hormone profiles and potentially changing the environment in which ovarian follicles develop. Because bovine granulosa cell culture is a commonly used technique and often bovine ovaries are collected from abattoirs with no record of implant status, the objective of this study was to determine if the presence of an implant during bovine granulosa cell development impacts follicle stimulating hormone-regulated steroidogenic enzyme expression. Paired ovaries were collected from 16 feedlot heifers subjected to 1 of 3 treatments:non-implanted (n=5), Revalor 200 for 28 d (n=5), or Revalor 200 for 84 d (n=6). Small follicle (1 to 5 mm) granulosa cells were isolated from each pair and incubated with phosphate buffered saline (n=16) or 100 ng/mL follicle stimulating hormone (n=16) for 24 h. Results:Granulosa cells of implanted heifers treated with follicle stimulating hormone produced medium concentrations of progesterone similar (P=0.22) to non-implanted heifers, while medium estradiol concentrations were increased (P<0.10) at 28 and 84 d compared to non-implanted heifers indicating efficacy of treatment. Additionally, real-time PCR analysis in response to follicle stimulating hormone treatment demonstrated a decrease in steroidogenic acute regulatory protein (P=0.05) mRNA expression in heifers implanted for 84 d and an increase in P450 side chain cleavage mRNA in granulosa cells of heifers implanted for 28 (P<0.10) or 84 d (P<0.05) compared to non-implanted females. However, no difference in expression of 3-beta-hydroxysteroid dehydrogenase (P=0.57) and aromatase (P=0.23) were demonstrated in implanted or non-implanted heifers. Conclusions:These results indicate follicles which develop in the presence of high concentrations of androgenic and estrogenic steroids via an implant tend to demonstrate an altered capacity to respond to follicle

  7. Effect of Supplemental Lutein and Zeaxanthin on Serum, Macular Pigmentation, and Visual Performance in Patients with Early Age-Related Macular Degeneration

    Directory of Open Access Journals (Sweden)

    Yang-Mu Huang

    2015-01-01

    Full Text Available Purpose. To compare the 2-year effect of multiple doses of lutein/zeaxanthin on serum, macular pigmentation, and visual performance on patients with early age-related macular degeneration (AMD. Methods. In this randomized, double-blinded, and placebo-controlled trial, 112 early AMD patients randomly received either 10 mg lutein, 20 mg lutein, a combination of lutein (10 mg and zeaxanthin (10 mg, or placebo daily for 2 years. Serum concentration of lutein/zeaxanthin, macular pigment optical density (MPOD, visual functions including best-spectacle corrected visual acuity (BCVA, contrast sensitivity (CS, flash recovery time (FRT, and vision-related quality of life (VFQ25 was quantified. Results. Serum lutein concentration and MPOD significantly increased in all the active treatment groups. Supplementation with 20 mg lutein was the most effective in increasing MPOD and CS at 3 cycles/degree for the first 48 weeks. However, they both significantly increased to the same peak value following supplementation with either 10 mg or 20 mg lutein during the intervention. No statistical changes of BCVA or FRT were observed during the trial. Conclusions. Long-term lutein supplementation could increase serum lutein concentration, MPOD, and visual sensitivities of early AMD patients. 10 mg lutein daily might be an advisable long-term dosage for early AMD treatment.

  8. The mare as a model for luteinized unruptured follicle syndrome: intrafollicular endocrine milieu.

    Science.gov (United States)

    Bashir, S T; Gastal, M O; Tazawa, S P; Tarso, S G S; Hales, D B; Cuervo-Arango, J; Baerwald, A R; Gastal, E L

    2016-03-01

    Luteinized unruptured follicle (LUF) syndrome is a recurrent anovulatory dysfunction that affects up to 23% of women with normal menstrual cycles and up to 73% with endometriosis. Mechanisms underlying the development of LUF syndrome in mares were studied to provide a potential model for human anovulation. The effect of extended increase in circulating LH achieved by administration of recombinant equine LH (reLH) or a short surge of LH and decrease in progesterone induced by prostaglandin F2α (PGF2α) on LUF formation (Experiment 1), identification of an optimal dose of COX-2 inhibitor (flunixin meglumine, FM; to block the effect of prostaglandins) for inducing LUFs (Experiment 2), and evaluation of intrafollicular endocrine milieu in LUFs (Experiment 3) were investigated. In Experiment 1, mares were treated with reLH from Day 7 to Day 15 (Day 0=ovulation), PGF2α on Day 7, or in combination. In Experiment 2, FM at doses of 2.0 or 3.0 mg/kg every 12 h and human chorionic gonadotropin (hCG) (1500 IU) were administered after a follicle ≥32 mm was detected. In Experiment 3, FM at a dose of 2.0 mg/kg every 12 h plus hCG was used to induce LUFs and investigate the intrafollicular endocrine milieu. No LUFs were induced by reLH or PGF2α treatment; however, LUFs were induced in 100% of mares using FM. Intrafollicular PGF2α metabolite, PGF2α, and PGE2 were lower and the ratio of PGE2:PGF2α was higher in the induced LUF group. Higher levels of intrafollicular E2 and total primary sex steroids were observed in the induced LUF group along with a tendency for higher levels of GH, cortisol, and T; however, LH, PRL, VEGF-A, and NO did not differ between groups. In conclusion, this study reveals part of the intrafollicular endocrine milieu and the association of prostaglandins in LUF formation, and indicates that the mare might be an appropriate model for studying the poorly understood LUF syndrome.

  9. MRI findings of bilateral juvenile granulosa cell tumor of the testis in a newborn presenting as intraabdominal masses

    Energy Technology Data Exchange (ETDEWEB)

    Yikilmaz, Ali [Erciyes Medical School, Department of Radiology, Gevher Nesibe Hospital, Kayseri (Turkey); Lee, Edward Y. [Children' s Hospital Boston and Harvard Medical School, Department of Radiology, Boston, MA (United States)

    2007-10-15

    Juvenile granulosa cell tumor (JGCT) of the testis is a rare benign tumor that typically presents as a relatively small (<2 cm) unilateral scrotal mass in neonates or infants. Bilateral JGCT of the testes presenting as large intraabdominal masses in the neonate is very rare. Utilizing preoperative MRI findings, we report a rare case of bilateral JGCT of the testes presenting as large multiseptated abdominal masses originating from undescended intraabdominal testes in a neonate. (orig.)

  10. Specific disruption of Tsc1 in ovarian granulosa cells promotes ovulation and causes progressive accumulation of corpora lutea.

    Directory of Open Access Journals (Sweden)

    Lin Huang

    Full Text Available Tuberous sclerosis complex 1 (Tsc1 is a tumor suppressor negatively regulating mammalian target of rapamycin complex 1 (mTORC1. It is reported that mice lacking Tsc1 gene in oocytes show depletion of primordial follicles, resulting in premature ovarian failure and subsequent infertility. A recent study indicated that deletion of Tsc1 in somatic cells of the reproductive tract caused infertility of female mice. However, it is not known whether specific disruption of Tsc1 in granulosa cells influences the reproductive activity of female mice. To clarify this problem, we mated Tsc1(flox/flox mice with transgenic mice strain expressing cyp19-cre which exclusively expresses in granulosa cells of the ovary. Our results demonstrated that Tsc1(flox/flox; cyp19-cre mutant mice were fertile, ovulating more oocytes and giving birth to more pups than control Tsc1(flox/flox mice. Progressive accumulation of corpora lutea occurred in the Tsc1(flox/flox; cyp19-cre mutant mice with advanced age. These phenotypes could be explained by the elevated activity of mTORC1, as indicated by increased phosphorylation of rpS6, a substrate of S6 in the Tsc1(flox/flox; cyp19-cre mutant granulosa cells. In addition, rapamycin, a specific mTORC1 inhibitor, effectively rescued the phenotype caused by increased mTORC1 activity in the Tsc1(cko ovaries. Our data suggest that conditional knockout of Tsc1 in granulosa cells promotes reproductive activity in mice.

  11. Mimosine As Well As Serum Starvation Can Be Used for Cell Cycle Synchronization of Sheep Granulosa Cells

    OpenAIRE

    2014-01-01

    This study was evaluated the effect of different synchronization protocols such as serum starvation for 1–3 days, confluency and chemical inhibitors on synchronization accuracy at G0/G1, apoptosis, and DNA synthesis in sheep granulosa cells. The cells were obtained from ovarian antral follicles of slaughtered sheep and used at first and fifth passages. Flow cytometry analysis showed that confluent cells, serum starvation for 24, 48, and 72 hours, and mimosine treatment significantly increase...

  12. Cells with Stem Cell Characteristics in Somatic Compartments of the Ovary

    Directory of Open Access Journals (Sweden)

    Katarzyna Kossowska-Tomaszczuk

    2013-01-01

    Full Text Available Antral follicular growth in the ovary is characterized by rapid expansion of granulosa cells accompanied by a rising complexity of their functionality. Within two weeks the number of human granulosa cells increases from less than 500,000 to more than 50 millions cells per follicle and differentiates into groups of cells with a variety of specialized functions involved in steroidogenesis, nursing the oocyte, and forming a functional syncitium. Both the rapid proliferation and different specialized functions of the granulosa cells can only be explained through the involvement of stem cells. However, luteinizing granulosa cells were believed to be terminally differentiated cells. Only recently, stem and progenitor cells with FSH-receptor activity were identified in populations of luteinizing granulosa cells obtained during oocyte collected for assisted reproduction. In the presence of the leukaemia-inhibiting factor (LIF, it was possible to culture a subpopulation of the luteinizing granulosa cells over prolonged time periods. Furthermore, when embedded in a matrix consisting of collagen type I, these cells continued to express the FSH receptor over prolonged time periods, developed globular formations that surrogated as follicle-like structures, providing a promising tool for reproductive biology.

  13. Active immunization to luteinizing hormone releasing hormone to inhibit the induction of mammary tumors in the rat

    Energy Technology Data Exchange (ETDEWEB)

    Ravdin, P.M.; Jordan, V.C.

    1988-01-01

    Immunization of female rats with a bovine serum albumin-luteinizing hormone releasing hormone conjugate results in suppression of dimethylbenzanthracene mammary tumor incidence. Tumor incidence was 1.3, and 1.29 tumors per rat in bovine serum albumin alone (n = 10) and unimmunized (n = 18) control groups, but no tumors were found in the bovine serum albumin-luteinizing hormone releasing hormone conjugate immunized animals (n = 10). In a second experiment immunization with bovine serum albumin-luteinizing hormone releasing hormone conjugates reduced tumor incidence to 0.3 tumors per rat (n = 10) from the 1.2 tumors per animal seen in the control animals (n = 10) immunized with bovine serum albumin alone. Bovine serum albumin-luteinizing hormone immunization caused the production of anti-LHRH antibodies, an interruption of estrous cycles, lowered serum estradiol and progesterone levels, and atrophy of the ovaries and uteri. Immunization BSA-hormone conjugates is a novel anti-tumor strategy.

  14. Modulation of Steroidogenic Pathway in Rat Granulosa Cells with Subclinical Cd Exposure and Insulin Resistance: An Impact on Female Fertility

    Directory of Open Access Journals (Sweden)

    Muskaan Belani

    2014-01-01

    Full Text Available Changes in lifestyle lead to insulin resistance (IR in females ultimately predisposing them towards infertility. In addition, cadmium (Cd, an environmental endocrine disruptor, is reported for detrimental effects on granulosa cells, thus leading to ovarian dysfunction. A combination of these factors, lifestyle and environment, seems to play a role in etiology of idiopathic infertility that accounts for 50% amongst the total infertility cases. To address this issue, we made an attempt to investigate the extent of Cd impact on insulin-resistant (IR granulosa cells. We exposed adult female Charles Foster rats to dexamethasone and confirmed IR condition by fasting insulin resistance index (FIRI. On treatment of IR rats with Cd, the preliminary studies demonstrated prolonged estrous cyclicity, decrease in serum estradiol concentrations, abnormal histology of ovary, and increased granulosa cell death. Further gene and protein expression studies of steroidogenic acute regulatory (StAR protein, 17β-hydroxysteroid dehydrogenase (17β-HSD, and cytochrome P450 aromatase (CYP19A1 were performed. Protein expression studies demonstrated significant decrease in treated groups when compared with control. Study revealed that, in spite of the molecular parameters being affected at varied level, overall ovarian physiology is maximally affected in IR and Cd coexposed group, thus mimicking the condition similar to those prevailing in infertile females.

  15. Apoptosis in Granulosa cells during follicular atresia:relationship with steroids and insulin-like growth factors

    Institute of Scientific and Technical Information of China (English)

    Yuan Song YU; Hong Shu SUI; Zheng Bin HAN; Wei LI; Ming Jiu LUO; Jing He TAN

    2004-01-01

    It is well known that during mammalian ovarian follicular development, the majority of follicles undergo atresia at various stages of their development. However, the mechanisms controlling this selection process remain unknown. In this study, we investigated apoptosis in granulosa cells during goat follicular atresia by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL). The changes in the levels of steroids, insulin-like growth factors (IGFs) and IGF receptors were studied by radioimmunoassay (RIA) and semi-quantitative reverse transcription-PCR. We found that the percentage of apoptotic granulosa cells in the atretic (A) follicles was significantly higher than that in the slightly atretic (SA) and healthy (H) follicles. The level of estradiol and the ratio of estradiol to progesterone in H follicles were significantly higher than those in A follicles. On the other hand, the level of progesterone was not significantly different among these follicle types. We also found that the level of IGF-I in H follicles was higher than in SA and A follicles, whereas the amount of IGF-Ⅱ did not vary significantly. The expression of IGF receptor also decreased in A follicles as compared to that in H and SA follicles. These results suggested that estradiol and IGF-I might be involved in controlling apoptosis in granulosa cells during follicular atresia.

  16. Effects of ergocryptine on plasma prolactin, luteinizing hormone, and progesterone in the periparturient sow.

    Science.gov (United States)

    Whitacre, M D; Threlfall, W R

    1981-09-01

    Effects of ergocryptine (ERG) on the periparturient sow were assessed by measuring changes in prolactin, luteinizing hormone, and progesterone in plasma. Hormones in plasma were determined on samples collected daily from 21 days before parturition until postpartum day (PPD) 21. Administration of ERG completely blocked the initiation of lactation and prevented the surge in prolactin that occurred just before parturition, as compared with effects in control sows. The gestation period was shorter (P less than 0.01) in ERG-treated sows compared with the gestation period in controls. Although the number of pigs farrowed was not different between treated and control sows, the ERG-treated sows did not raise any pigs to PPD 21. Luteinizing hormone concentrations were higher (P less than 0.01) from 10 days before parturition until PPD 21, but plasma progesterone concentrations were not different in treated sows when compared with concentrations in control sows.

  17. Regulation of gene expression in ovarian cancer cells by luteinizing hormone receptor expression and activation

    Directory of Open Access Journals (Sweden)

    Dam Phuongan

    2011-06-01

    Full Text Available Abstract Background Since a substantial percentage of ovarian cancers express gonadotropin receptors and are responsive to the relatively high concentrations of pituitary gonadotropins during the postmenopausal years, it has been suggested that receptor activation may contribute to the etiology and/or progression of the neoplasm. The goal of the present study was to develop a cell model to determine the impact of luteinizing hormone (LH receptor (LHR expression and LH-mediated LHR activation on gene expression and thus obtain insights into the mechanism of gonadotropin action on ovarian surface epithelial (OSE carcinoma cells. Methods The human ovarian cancer cell line, SKOV-3, was stably transfected to express functional LHR and incubated with LH for various periods of time (0-20 hours. Transcriptomic profiling was performed on these cells to identify LHR expression/activation-dependent changes in gene expression levels and pathways by microarray and qRT-PCR analyses. Results Through comparative analysis on the LHR-transfected SKOV-3 cells exposed to LH, we observed the differential expression of 1,783 genes in response to LH treatment, among which five significant families were enriched, including those of growth factors, translation regulators, transporters, G-protein coupled receptors, and ligand-dependent nuclear receptors. The most highly induced early and intermediate responses were found to occupy a network impacting transcriptional regulation, cell growth, apoptosis, and multiple signaling transductions, giving indications of LH-induced apoptosis and cell growth inhibition through the significant changes in, for example, tumor necrosis factor, Jun and many others, supportive of the observed cell growth reduction in in vitro assays. However, other observations, e.g. the substantial up-regulation of the genes encoding the endothelin-1 subtype A receptor, stromal cell-derived factor 1, and insulin-like growth factor II, all of which are

  18. Regulation of endometrial cancer cell growth by luteinizing hormone (LH) and follicle stimulating hormone (FSH)

    OpenAIRE

    Davies, S.; Bax, C M R; Chatzaki, E; Chard, Tim; Iles, Ray K.

    2000-01-01

    Gonadotrophin releasing hormone analogues (GnRHa) have been used to treat recurrent endometrial cancer. However, the mode of action is uncertain. Our previous studies showed no direct effect of GnRHa on endometrial cancer cell growth in vitro. We have now examined the effect of luteinizing hormone (LH) and follicle stimulating hormone (FSH) on endometrial cancer cell growth. The aim was to determine whether suppression of pituitary LH and FSH by GnRHa could explain the tumour regression seen ...

  19. Metabolic alterations of lutein, β-carotene and chlorophyll a during germination of two soybean sprout varieties.

    Science.gov (United States)

    Lee, Jinwook; Hwang, Young-Sun; Lee, Jeong-Dong; Chang, Woo-Suk; Choung, Myoung-Gun

    2013-12-01

    The metabolic changes of lutein, β-carotene and chlorophyll a during germination of the soybean (Glycine max (L.) Merr.) sprout varieties, 'Pungsannamulkong' and 'Bosug', have been studied. Seeds were germinated at 20 °C with 80% humidity in the darkness and sampled at 2 day intervals for 10 days. Partial least squares (PLS) scores plot showed that the responses of three metabolites during germination were linearly linked with each other except for day 2 in both varieties. PLS loading plots indicated that lutein content in whole sprout and cotyledon was closely associated with germination in 'Pungsannamulkong' while the chlorophyll a content in whole sprouts was highly linked with germination in 'Bosug'. Heatmap analyses revealed that lutein and β-carotene levels, but not those of chlorophyll a, accumulated in whole soybean sprouts and cotyledon. While hypocotyls did not accumulate lipophilic pigments during germination, the accumulation of lutein and β-carotene in the cotyledons was greater in 'Pungsannamulkong' than in 'Bosug' sprouts. In addition, the contents of lutein, β-carotene and chlorophyll a increased from those in the seeds. Overall, the metabolic changes of lutein, β-carotene and chlorophyll a during germination are affected not only by variety but also by organ type.

  20. A single extraction and HPLC procedure for simultaneous analysis of phytosterols, tocopherols and lutein in soybeans.

    Science.gov (United States)

    Slavin, Margaret; Yu, Liangli Lucy

    2012-12-15

    A saponification/extraction procedure and high performance liquid chromatography (HPLC) analysis method were developed and validated for simultaneous analysis of phytosterols, tocopherols and lutein (a carotenoid) in soybeans. Separation was achieved on a phenyl column with a ternary, isocratic solvent system of acetonitrile, methanol and water (48:22.5:29.5, v/v/v). Evaporative light scattering detection (ELSD) was used to quantify β-sitosterol, stigmasterol, campesterol, and α-, δ- and γ-tocopherols, while lutein was quantified with visible light absorption at 450 nm. Peak identification was verified by retention times and spikes with external standards. Standard curves were constructed (R(2)>0.99) to allow for sample quantification. Recovery of the saponification and extraction was demonstrated via analysis of spiked samples. Also, the accuracy of results of four soybeans using the described saponification and HPLC analytical method was validated against existing methods. This method offers a more efficient alternative to individual methods for quantifying lutein, tocopherols and sterols in soybeans.

  1. Lutein, Zeaxanthin, and meso-Zeaxanthin in the Clinical Management of Eye Disease.

    Science.gov (United States)

    Scripsema, Nicole K; Hu, Dan-Ning; Rosen, Richard B

    2015-01-01

    Lutein, zeaxanthin, and meso-zeaxanthin are xanthophyll carotenoids found within the retina and throughout the visual system. The retina is one of the most metabolically active tissues in the body. The highest concentration of xanthophylls is found within the retina, and this selective presence has generated many theories regarding their role in supporting retinal function. Subsequently, the effect of xanthophylls in the prevention and treatment of various eye diseases has been examined through epidemiological studies, animal studies, and clinical trials. This paper attempts to review the epidemiological studies and clinical trials investigating the effects of xanthophylls on the incidence and progression of various eye diseases. Observational studies have reported that increased dietary intake and higher serum levels of lutein and zeaxanthin are associated with lower risk of age-related macular degeneration (AMD), especially late AMD. Randomized, placebo-controlled clinical trials have demonstrated that xanthophyll supplementation increases macular pigment levels, improves visual function, and decreases the risk of progression to late AMD, especially neovascular AMD. Current publications on the preventive and therapeutic effects of lutein and zeaxanthin on cataracts, diabetic retinopathy, and retinopathy of prematurity have reported encouraging results.

  2. The effects of levonorgestrel on FSH-stimulated primary rat granulosa cell cultures through gene expression profiling are associated to hormone and folliculogenesis processes.

    Science.gov (United States)

    Lira-Albarrán, Saúl; Larrea-Schiavon, Marco F; González, Leticia; Durand, Marta; Rangel, Claudia; Larrea, Fernando

    2017-01-05

    Levonorgestrel (LNG), a synthetic progestin, is used in emergency contraception (EC). The mechanism is preventing or delaying ovulation at the level of the hypothalamic pituitary unit; however, little knowledge exists on LNG effects at the ovary. The aim of this study was to identify the effects of LNG on FSH-induced 17β-estradiol (E2) production, including LNG-mediated changes on global gene expression in rat granulosa cells (GC). Isolated GC from female Wistar rats were incubated in vitro in the presence or absence of human FSH and progestins. At the end of incubations, culture media and cells were collected for E2 and mRNA quantitation. The results showed the ability of LNG to inhibit both hFSH-induced E2 production and aromatase gene expression. Microarray analysis revealed that LNG treatment affects GC functionality particularly that related to folliculogenesis and steroid metabolism. These results may offer additional evidence for the mechanisms of action of LNG as EC. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  3. Regulatory effect of hypoxia-inducible factor-1α on hCG-stimulated endothelin-2 expression in granulosa cells from the PMSG-treated rat ovary.

    Science.gov (United States)

    Zhang, Jisen; Zhang, Zhenghong; Wu, Yanqing; Chen, Liyun; Luo, Qianping; Chen, Jiajie; Huang, Xiaohong; Cheng, Yong; Wang, Zhengchao

    2012-01-01

    Endothelin (ET)-2 plays a crucial role in ovarian ovulation in mammals. The present study was designed to test the hypothesis that hypoxia-inducible factor (HIF)-1α-mediated transcriptional activation contributes to the increased expression of ET-2 gene in response to hCG in rat ovarian granulosa cells (GCs) during gonadotropin-induced superovulation. By real-time RT-PCR analysis, ET-2 mRNA expression was found to significantly increase in cultured ovarian GCs after treatment with hCG, or even N-carbobenzoxyl-L-leucinyl-L-leucinyl-L-norvalinal (MG-132), while this increased ET-2 mRNA expression could also be blocked by ferrous ammonium sulfate (FAS) under human chorionic gonadotropin (hCG) treatment. Further analysis also found that these changes of ET-2 mRNA were consistent with HIF-1α expression or HIF-1 activity, and HIF-1α inhibitor echinomycin inhibited ovulation in rats. Taken together, these results indicate that ET-2 is transcriptionally activated by hCG through HIF-1α-mediated mechanism in GCs. This HIF-1α-induced transcriptional activation may be one of the important mechanisms mediating the increase of ET-2 expression in GCs during the gonadotropin-induced mammalian ovulatory process in vivo.

  4. LH-receptor gene expression in human granulosa and cumulus cells from antral and preovulatory follicles

    DEFF Research Database (Denmark)

    Jeppesen, Janni Vikkelsø; Kristensen, Stine Gry; Nielsen, Maria Eilsø

    2012-01-01

    frozen and patients undergoing infertility treatment. Interventions: Cells and fluids were isolated from surgically excised ovaries or from aspirated preovulatory follicles. Main Outcome Measures: We evaluated gene expression of LHR, FSHR, androgen receptor (AR), aromatase (CYP19a1), and AMHR2 normalized...... to the GAPDH expression and associated with FF levels of anti-Mullerian hormone, inhibin-B, and steroids. Results: LHR expression was maximal in GC from preovulatory follicles before ovulation induction. A majority of 150 antral follicles (3-10 mm in diameter) showed LHR expression at approximately 10...

  5. The effect of recombinant human luteinizing hormone supplementation on IVF/ICSI outcome in women with ovarian hypofunction——aretrospective study in the GnRH antagonist protocol%促性腺激素释放素拮抗剂联合人重组促黄体激素对卵巢功能减退患者妊娠结局的影响

    Institute of Scientific and Technical Information of China (English)

    刁飞扬; 王嫜; 丁卫; 冒韵东; 刘嘉茵

    2011-01-01

    目的:研究重组人促黄体激素(r-hLH)在促性腺激素释放素(GnRH)拮抗剂促排卵方案中的应用指征.方法:回顾性分析2007年8月至2010年8月在本中心以GnRH拮抗剂方案促排卵的体外受精(IVF/ICSI)患者.卵巢功能减退纳入标准为卵泡刺激数(FSH)≥10U/L或窦卵泡数≤6个.研究分为两部分,第一部分纳入卵巢功能减退患者共107例,A组不添加r-hLH(n=80),B组添加r-hLH 75 U/d(n= 27).第二部分纳入应用GnRH拮抗剂+r-hLH+GnRH激动剂扳机的患者共71例并分为两组,C组为卵巢功能正常组(n=47),D组为卵巢功能减退组(n=24).评价两组卵巢反应和妊娠结局的临床指标包括成熟卵子数、受精率、卵裂率、优质胚胎率、种植率和临床妊娠率.结果:①卵巢功能减退患者中,添加r-hLH组每优势卵泡平均雌激素水平、IVF成熟卵子数(100.00% vs 96.31%)显著高于对照组,种植率(29.41% vs 19.46%)和临床妊娠率(44.44% vs31.25%)均高于对照组,但差异无显著性.②添加了r-hLH的患者中,卵巢功能减退组IVF成熟卵子数(100.00% vs 93.71%)、IVF受精率(79.52% vs 38.46%)和ICSI受精率(84.85% vs 42.74%)显著高于卵巢功能正常组,种植率(28.89% vs 17.44%)、临床妊娠率(41.67%vs 25.53%)均高于卵巢功能正常组,但差异无显著性.结论:在GnRH拮抗剂方案中,添加r-hLH可以改善卵巢反应减退患者的卵巢反应和临床结局.卵巢功能正常者不是添加r-hLH的适应人群.%Objective: To study the indication of human recombinant luteinizing hormone (r-hLH) in GnRH antagonist Methods: Retrospective analysis. All the patients using GnRH antagonist protocol were enrolled in this study from Aug,200 2010. This study was divided into two parts. (D The first part enrolled 107 patients with ovarian hypofunction (FSH^ 10U/I follicle count ^6) ,80 of them without supplementation of r-hLH (group A) ,27 of them received r-hLH together with GnRH , (group B). ? The second part enrolled

  6. A quantitative histochemical study of lactate dehydrogenase and succinate dehydrogenase activities in the membrana granulosa of the ovulatory follicle of the rat.

    Science.gov (United States)

    Zoller, L C; Enelow, R

    1983-11-01

    Using a microdensitometer, lactate dehydrogenase and succinate dehydrogenase activities were measured in the membrana granulosa of the rat ovulatory follicle. Ovaries were removed on each day of the oestrous cycle; oestrus, dioestrus-1, dioestrus-2, and proestrus; and enzyme activities measured in the membrana granulosa as a whole and in four regions within it: peripheral (PR), antral (AR), cumulus oophorus (CO) and corona radiata (CR). Throughout the cycle, lactate dehydrogenase activity was greatest in PR. On oestrus, lactate dehydrogenase activity was progressively less in AR, CO and CR. On dioestrus-1, activity was identical in AR and CO and less in CR. On dioestrus-2, activity was greater in AR than in CO or CR. By proestrus, activity was equal in AR, CO and CR. In the membrana granulosa as a whole, and in each region, lactate dehydrogenase activity declined as ovulation approached. In contrast, succinate dehydrogenase activity in the membrana granulosa as a whole and in PR was constant throughout the cycle. Activity fluctuated in the other regions. Succinate dehydrogenase activity on oestrus was greatest in PR, less in AR and CO and least in CR. On the remaining days, succinate dehydrogenase activity was greatest in PR and less but equal in the remainder of the membrana granulosa.

  7. Participation of Mitogen-activated Protein Kinase in Luteinizing Hormone-induced Differential Regulation of Steroidogenesis and Steroidogenic Gene Expression in Mural and Cumulus Granulosa Cells of Mouse Preovulatory Follicles

    DEFF Research Database (Denmark)

    Su, You-Qiang; Nyegaard, Mette; Overgaard, Michael Toft

    2006-01-01

    superovulation model for premature female mice was used to obtain MGCs and cumulus-oocyte complexes (COCs), and sensitive real-time reverse transcription polymerase chain reaction was used to simultaneously detect the expression levels of transcripts encoding key steroidogenic enzymes in the same sample. We...

  8. Identification of Polycystic Ovary Syndrome (PCOS) Specific Genes in Cumulus and Mural Granulosa Cells

    Science.gov (United States)

    Aydos, Alp; Gurel, Aykut; Oztemur Islakoglu, Yasemin; Noyan, Senem; Gokce, Bagdagul; Ecemis, Tolga; Kaya, Cemil; Aksu, Arif Tarik

    2016-01-01

    Polycystic ovary syndrome (PCOS) is a metabolic and endocrine disorder which affects women of reproductive age with prevalence of 8–18%. The oocyte within the follicle is surrounded by cumulus cells (CCs), which connect with mural granulosa cells (MGCs) that are responsible for secreting steroid hormones. The main aim of this study is comparing gene expression profiles of MGCs and CCs in PCOS and control samples to identify PCOS-specific differentially expressed genes (DEGs). In this study, two microarray databases were searched for mRNA expression microarray studies performed with CCs and MGCs obtained from PCOS patients and control samples. Three independent studies were selected to be integrated with naive meta-analysis since raw meta-data from these studies were found to be highly correlated. DEGs in these somatic cells were identified for PCOS and control groups. This study enabled us to reveal dysregulation in MAPK (mitogen activated protein kinase), insulin and Wnt signaling pathways between CCs and MGCs in PCOS. The meta-analysis results together with qRT-PCR validations provide evidence that molecular signaling is dysregulated through MGCs and CCs in PCOS, which is important for follicle and oocyte maturation and may contribute to the pathogenesis of the syndrome. PMID:27997581

  9. Tumor de células da granulosa com metástases numa gata

    Directory of Open Access Journals (Sweden)

    Márcia Regina Giacóia

    1999-01-01

    Full Text Available As características clínicas, macroscópicas e microscópicas de um tumor de células da granulosa no ovário de uma gata de 2 anos de idade são descritas neste trabalho. Essa neoplasia é rara em gatos, principalmente devido à idade apresentada pelo animal. O diagnóstico foi feito clinicamente através de exame ultra-sonográfico. O tumor, uma massa cística na região do ovário esquerdo, metastatizou para o epíploo e para os pulmões. As células tumorais expressaram vimentina e arranjavam-se num padrão sarcomatoso e difuso, sustentado por fino estroma fibrovascular. A presença de sinais clínicos, como perda de pêlos e repetição de estros, é indicativa de síndrome paraneoplásica ortoendócrina, devido à excessiva estimulação estrogênica. A existência desse tipo de tumor deve ser considerada no diagnóstico diferencial de distúrbios comportamentais em gatas.

  10. Low oxygen level increases proliferation and metabolic changes in bovine granulosa cells.

    Science.gov (United States)

    Shiratsuki, Shogo; Hara, Tomotaka; Munakata, Yasuhisa; Shirasuna, Koumei; Kuwayama, Takehito; Iwata, Hisataka

    2016-12-05

    The present study addresses molecular backgrounds underlying low oxygen induced metabolic changes and 1.2-fold change in bovine granulosa cell (GCs) proliferation. RNA-seq revealed that low oxygen (5%) upregulated genes associated with HIF-1 and glycolysis and downregulated genes associated with mitochondrial respiration than that in high oxygen level (21%). Low oxygen level induced high glycolytic activity and low mitochondrial function and biogenesis. Low oxygen level enhanced GC proliferation with high expression levels of HIF-1, VEGF, AKT, mTOR, and S6RP, whereas addition of anti-VEGF antibody decreased cellular proliferation with low phosphorylated AKT and mTOR expression levels. Low oxygen level reduced SIRT1, whereas activation of SIRT1 by resveratrol increased mitochondrial replication and decreased cellular proliferation with reduction of phosphorylated mTOR. These results suggest that low oxygen level stimulates the HIF1-VEGF-AKT-mTOR pathway and up-regulates glycolysis, which contributes to GC proliferation, and downregulation of SIRT1 contributes to hypoxia-associated reduction of mitochondria and cellular proliferation.

  11. MicroRNA Mediating Networks in Granulosa Cells Associated with Ovarian Follicular Development

    Science.gov (United States)

    Zhang, Baoyun; Chen, Long; Feng, Guangde; Xiang, Wei; Zhang, Ke; Chu, Mingxing

    2017-01-01

    Ovaries, which provide a place for follicular development and oocyte maturation, are important organs in female mammals. Follicular development is complicated physiological progress mediated by various regulatory factors including microRNAs (miRNAs). To demonstrate the role of miRNAs in follicular development, this study analyzed the expression patterns of miRNAs in granulosa cells through investigating three previous datasets generated by Illumina miRNA deep sequencing. Furthermore, via bioinformatic analyses, we dissected the associated functional networks of the observed significant miRNAs, in terms of interacting with signal pathways and transcription factors. During the growth and selection of dominant follicles, 15 dysregulated miRNAs and 139 associated pathways were screened out. In comparison of different styles of follicles, 7 commonly abundant miRNAs and 195 pathways, as well as 10 differentially expressed miRNAs and 117 pathways in dominant follicles in comparison with subordinate follicles, were collected. Furthermore, SMAD2 was identified as a hub factor in regulating follicular development. The regulation of miR-26a/b on smad2 messenger RNA has been further testified by real time PCR. In conclusion, we established functional networks which play critical roles in follicular development including pivotal miRNAs, pathways, and transcription factors, which contributed to the further investigation about miRNAs associated with mammalian follicular development.

  12. Loss of inhibin alpha uncouples oocyte-granulosa dynamics and disrupts postnatal folliculogenesis

    Science.gov (United States)

    Myers, Michelle; Middlebrook, Brooke S.; Matzuk, Martin M.; Pangas, Stephanie A.

    2009-01-01

    Targeted disruption of the inhibin α gene (Inha−/−) in mice results in an ovarian phenotype of granulosa cell tumors that renders the animals infertile. Little is known about the reproductive defects prior to tumor development. Here, we report novel data on early follicle dynamics in Inha−/− mice, which demonstrate that inhibin α has important consequences upon follicle development. Morphological changes in both germ and somatic cells were evident in postnatal day 12 ovaries, with Inha−/− mice exhibiting numerous multilayered follicles that were far more advanced than those observed in age-matched controls. These changes were accompanied by alterations in follicle dynamics such that Inha−/− ovaries had fewer follicles in the resting pool and more committed in the growth phase. Absence of inhibin α resulted in advanced follicular maturation as marked by premature loss of anti-Müllerian hormone (AMH) in secondary follicles. Additionally, gene expression analysis revealed changes in factors known to be vital for oocyte and follicle development. Together, these data provide key evidence to suggest that regulation of the inhibin/activin system is essential for early folliculogenesis in the prepubertal mouse ovary. PMID:19666016

  13. Moxibustion Reduces Ovarian Granulosa Cell Apoptosis Associated with Perimenopause in a Natural Aging Rat Model

    Directory of Open Access Journals (Sweden)

    Xiao-Lan Shi

    2015-01-01

    Full Text Available In recent years, concerns about the adverse effects of hormone replacement therapy have increased interest in alternative therapies for the management of the symptoms of perimenopause. Here, we investigated the effects of moxibustion, a traditional Chinese practice that is involved in heated Artemisia vulgaris (mugwort stimulation, on hormonal imbalance and ovarian granulosa cell (GC apoptosis in a rat model of perimenopause. Our results showed that mild warm moxibustion (MWM modulated the circulating levels of estradiol and follicle-stimulating hormone and their receptors and inhibited apoptosis in the ovaries of perimenopausal rats, similar to the effect of estrogen. Further investigation revealed that the effects of MWM on ovary tissues and cultured GCs were mediated by the modulation of the activity of Forkhead box protein O1 and involved the JAK2/STAT3 pathway. Our results provide information on the factors and pathways modulated by MWM and shed light on the mechanism underlying the beneficial effect of moxibustion on the symptoms of perimenopause.

  14. Female newts (Taricha granulosa) produce tetrodotoxin laden eggs after long term captivity.

    Science.gov (United States)

    Gall, Brian G; Stokes, Amber N; French, Susannah S; Brodie, Edmund D; Brodie, Edmund D

    2012-11-01

    We investigated the presence of tetrodotoxin (TTX) in the eggs of wild-caught newts (Taricha granulosa) at capture and again after one, two, and three years in captivity. Females initially produced eggs that contained quantities of TTX similar to previous descriptions of eggs from wild-caught adults. After the first year in captivity, the egg toxicity from each female declined, ultimately remaining constant during each of the successive years in captivity. Despite declining, all females continued to produce eggs containing substantial quantities of TTX during captivity. The decline in toxicity can not be attributed to declining egg mass but may be the result of the abbreviated reproductive cycle to which the captive newts were subjected in the lab. Finally, an estimate of the amount of TTX provisioned in the entire clutch from each female is similar to the quantity of TTX regenerated in the skin after electrical stimulation. These results, coupled with other long-term studies on the maintenance and regeneration of TTX in the skin, suggests an endogenous origin of TTX in newts.

  15. STMN1 Promotes Progesterone Production Via StAR Up-regulation in Mouse Granulosa Cells

    Science.gov (United States)

    Dou, Yun-De; Zhao, Han; Huang, Tao; Zhao, Shi-Gang; Liu, Xiao-Man; Yu, Xiao-Chen; Ma, Zeng-Xiang; Zhang, Yu-Chao; Liu, Tao; Gao, Xuan; Li, Lei; Lu, Gang; Chan, Wai-Yee; Gao, Fei; Liu, Hong-Bin; Chen, Zi-Jiang

    2016-01-01

    Stathmin 1 (STMN1) is a biomarker in several types of neoplasms. It plays an important role in cell cycle progression, mitosis, signal transduction and cell migration. In ovaries, STMN1 is predominantly expressed in granulosa cells (GCs). However, little is known about the role of STMN1 in ovary. In this study, we demonstrated that STMN1 is overexpressed in GCs in patients with polycystic ovary syndrome (PCOS). In mouse primary GCs, the overexpression of STMN1 stimulated progesterone production, whereas knockdown of STMN1 decreased progesterone production. We also found that STMN1 positively regulates the expression of Star (steroidogenic acute regulatory protein) and Cyp11a1 (cytochrome P450 family 11 subfamily A member 1). Promoter and ChIP assays indicated that STMN1 increased the transcriptional activity of Star and Cyp11a1 by binding to their promoter regions. The data suggest that STMN1 mediates the progesterone production by modulating the promoter activity of Star and Cyp11a1. Together, our findings provide novel insights into the molecular mechanisms of STMN1 in ovary GC steroidogenesis. A better understanding of this potential interaction between STMN1 and Star in progesterone biosynthesis in GCs will facilitate the discovery of new therapeutic targets in PCOS. PMID:27270953

  16. Antioxidant status and selected biochemical parameters of porcine ovarian granulosa cells exposed to lead in vitro.

    Science.gov (United States)

    Capcarová, Marcela; Kolesárová, Adriana; Lukác, Norbert; Sirotkin, Alexander; Roychoudhury, Shubhadeep

    2009-12-01

    The objective of this study was to determine the activity of superoxide dismutase (SOD), total antioxidant status (TAS) and release of calcium, phosphorus, magnesium, sodium, potassium, total lipids, totals proteins, glucose, cholesterol and triglycerides by porcine ovarian granulosa cells cultured in vitro after lead acetate administration. The parameters were analyzed using semi-automated clinical chemistry analyzer Microlab 300, microprocessor-controlled analyzer EasyLite and spectrophotometer Genesys 10. Cells were cultured with lead acetate trihydrate [Pb(CH(3)COO)(2).3H(2)O] as follows: group Max (5 mg Pb(CH(3)COO)(2).3H(2)O/10 mL), group A (2.5 mg/10 mL), group B (0.83 mg/10 mL), group C (0.625 mg/10 mL), group D (0.455 mg/10 mL) and the control group without lead exposure for 18 hrs. The highest TAS was estimated in the control group without lead treatment in comparison with other groups (MAX, A, B, C, D). Statistical analyses showed significantly lower value (P 0.05) were detected in concentration of other studied parameters among observed groups, too.

  17. Plasma lutein and zeaxanthin and the risk of age-related nuclear cataract among the elderly Finnish population.

    Science.gov (United States)

    Karppi, Jouni; Laukkanen, Jari A; Kurl, Sudhir

    2012-07-14

    Oxidative stress plays an important role in cataractogenesis. Previous studies have shown that long-term dietary intake of antioxidants (lutein and zeaxanthin) may decrease the risk of age-related cataracts. The aim of the present study was to examine whether plasma concentrations of lutein and zeaxanthin are related to age-related nuclear cataract in the elderly population. Subjects were participants in the Kuopio Ischaemic Heart Disease Risk Factor Study and they were classified into tertiles according to plasma concentrations of lutein and zeaxanthin. The association of plasma lutein and zeaxanthin concentrations with age-related nuclear cataract in 1689 elderly subjects (aged 61-80 years) was investigated in the present cross-sectional study by using the Cox proportional hazards model. A total of 113 cases of incident age-related cataracts were confirmed, of which 108 cases were nuclear cataracts. After adjustment for age, examination year, sex, BMI, smoking, alcohol consumption, serum LDL-cholesterol, serum HDL-cholesterol, years of education, use of oral corticosteroids, history of diabetes and history of hypertension with current use of antihypertensive medication, subjects in the highest tertiles of plasma concentrations of lutein and zeaxanthin had 42 and 41 % lower risks of nuclear cataract, respectively, compared with those in the lowest tertiles (relative risk (RR) = 0·58, 95 % CI 0·35, 0·98; P = 0·041 for lutein and RR = 0·59, 95 % CI 0·35, 0·99; P = 0·046 for zeaxanthin). In conclusion, we suggest that high plasma concentrations of lutein and zeaxanthin were associated with a decreased risk of age-related nuclear cataract in the elderly population.

  18. Lutein protects against methotrexate-induced and reactive oxygen species-mediated apoptotic cell injury of IEC-6 cells.

    Science.gov (United States)

    Chang, Chi-Jen; Lin, Ji-Fan; Chang, Hsun-Hsien; Lee, Gon-Ann; Hung, Chi-Feng

    2013-01-01

    High-dose chemotherapy using methotrexate (MTX) frequently induces side effects such as mucositis that leads to intestinal damage and diarrhea. Several natural compounds have been demonstrated of their effectiveness in protecting intestinal epithelial cells from these adverse effects. In this paper, we investigated the protection mechanism of lutein against MTX-induced damage in IEC-6 cells originating from the rat jejunum crypt. The cell viability, induced-apoptosis, reactive oxygen species (ROS) generation, and mitochondrial membrane potential in IEC-6 cells under MTX treatment were examined in the presence or absence of lutein. Expression level of Bcl2, Bad and ROS scavenging enzymes (including SOD, catalase and Prdx1) were detected by quantitative RT-PCR. The cell viability of IEC-6 cells exposed to MTX was decreased in a dose- and time-dependent manner. MTX induces mitochondrial membrane potential loss, ROS generation and caspase 3 activation in IEC-6 cells. The cytotoxicity of MTX was reduced in IEC-6 cells by the 24 h pre-treatment of lutein. We found that pre-treatment of lutein significantly reduces MTX-induced ROS and apoptosis. The expression of SOD was up-regulated by the pre-treatment of lutein in the MTX-treated IEC-6 cells. These results indicated that lutein can protect IEC-6 cells from the chemo-drugs induced damage through increasing ROS scavenging ability. The MTX-induced apoptosis of IEC-6 cells was shown to be repressed by the pre-treatment of lutein, which may represent a promising adjunct to conventional chemotherapy for preventing intestinal damages.

  19. Highly potent analogues of luteinizing hormone-releasing hormone containing D-phenylalanine nitrogen mustard in position 6

    Energy Technology Data Exchange (ETDEWEB)

    Bajusz, S.; Janaky, T.; Csernus, V.J.; Bokser, L.; Fekete, M.; Srkalovic, G.; Redding, T.W.; Schally, A.V. (Tulane Univ. School of Medicine, New Orleans, LA (USA))

    1989-08-01

    The nitrogen mustard derivatives of 4-phenylbutyric acid and L-phenylalanine, called chlorambucil (Chl) and melphalan (Mel), respectively, have been incorporated into several peptide hormones, including luteinizing hormone-releasing hormone (LH-RH). The alkylating analogues of LH-RH were prepared by linking Chl, as an N-acyl moiety, to the complete amino acid sequence of agonistic and antagonistic analogues. These compounds, in particular the antagonistic analogues, showed much lower potency than their congeners carrying other acyl groups. To obtain highly potent alkylating analogues of LH-RH, the D enantiomer of Mel was incorporated into position 6 of the native hormone and some of its antagonistic analogues. Of the peptides prepared, (D-Mel{sup 6})LH-RH (SB-05) and (Ac-D-Nal(2){sup 1},D-Phe(pCl){sup 2},D-Pal(3){sup 3},Arg{sup 5},D-Mel{sup 6},D-Ala{sup 10})LH-RH (SB-86, where Nal(2) is 3-(2-naphthyl)alanine and Pal(3) is 3-(3-pyridyl)alanine) possessed the expected high agonistic and antagonistic activities, respectively, and also showed high affinities for the membrane receptors of rat pituitary cells, human breast cancer cells, human prostate cancer cells, and rat Dunning R-3327 prostate tumor cells. These two analogues exerted cytotoxic effects on human and rat mammary cancer cells in vitro. Thus these two D-Mel{sup 6} analogues seem to be particularly suitable for the study of how alkylating analogues of LH-RH could interfere with intracellular events in certain cancer cells.

  20. Lutein from Deepoxidation of Lutein Epoxide Replaces Zeaxanthin to Sustain an Enhanced Capacity for Nonphotochemical Chlorophyll Fluorescence Quenching in Avocado Shade Leaves in the Dark1

    Science.gov (United States)

    Förster, Britta; Pogson, Barry James; Osmond, Charles Barry

    2011-01-01

    Leaves of avocado (Persea americana) that develop and persist in deep shade canopies have very low rates of photosynthesis but contain high concentrations of lutein epoxide (Lx) that are partially deepoxidized to lutein (L) after 1 h of exposure to 120 to 350 μmol photons m−2 s−1, increasing the total L pool by 5% to 10% (ΔL). Deepoxidation of Lx to L was near stoichiometric and similar in kinetics to deepoxidation of violaxanthin (V) to antheraxanthin (A) and zeaxanthin (Z). Although the V pool was restored by epoxidation of A and Z overnight, the Lx pool was not. Depending on leaf age and pretreatment, the pool of ΔL persisted for up to 72 h in the dark. Metabolism of ΔL did not involve epoxidation to Lx. These contrasting kinetics enabled us to differentiate three states of the capacity for nonphotochemical chlorophyll fluorescence quenching (NPQ) in attached and detached leaves: ΔpH dependent (NPQΔpH) before deepoxidation; after deepoxidation in the presence of ΔL, A, and Z (NPQΔLAZ); and after epoxidation of A+Z but with residual ΔL (NPQΔL). The capacity of both NPQΔLAZ and NPQΔL was similar and 45% larger than NPQΔpH, but dark relaxation of NPQΔLAZ was slower. The enhanced capacity for NPQ was lost after metabolism of ΔL. The near equivalence of NPQΔLAZ and NPQΔL provides compelling evidence that the small dynamic pool ΔL replaces A+Z in avocado to “lock in” enhanced NPQ. The results are discussed in relation to data obtained with other Lx-rich species and in mutants of Arabidopsis (Arabidopsis thaliana) with increased L pools. PMID:21427278

  1. Dimeric ligands for GPCRs involved in human reproduction : synthesis and biological evaluation

    NARCIS (Netherlands)

    Bonger, Kimberly Michelle

    2008-01-01

    Dimeric ligands for G-protein coupled receptors that are involved in human reproduction, namely the gonadotropin releasing hormone receptor, the luteinizing hormone receptor and the follicle-stimulating hormone receptor, were synthesized and biologically evaluated.

  2. Granulosa Cell-Expressed BMPR1A and BMPR1B Have Unique Functions in Regulating Fertility but Act Redundantly to Suppress Ovarian Tumor Development

    Science.gov (United States)

    Edson, Mark A.; Nalam, Roopa L.; Clementi, Caterina; Franco, Heather L.; DeMayo, Francesco J.; Lyons, Karen M.; Pangas, Stephanie A.; Matzuk, Martin M.

    2010-01-01

    Bone morphogenetic proteins (BMPs) have diverse roles in development and reproduction. Although several BMPs are produced by oocytes, thecal cells, and granulosa cells of developing follicles, the in vivo functions of most of these ligands are unknown. BMP signals are transduced by multiple type I and type II TGFβ family receptors, and of the type I receptors, BMP receptor 1A (BMPR1A) and BMP receptor 1B (BMPR1B) are known to be expressed in rodent granulosa cells. Female mice homozygous null for Bmpr1b are sterile due to compromised cumulus expansion, but the function of BMPR1A in the ovary is unknown. To further decipher a role for BMP signaling in mouse granulosa cells, we deleted Bmpr1a in the granulosa cells of the ovary and found Bmpr1a conditional knockout females to be subfertile with reduced spontaneous ovulation. To explore the redundant functions of BMP receptor signaling in the ovary, we generated Bmpr1a Bmpr1b double-mutant mice, which developed granulosa cell tumors that have evidence of increased TGFβ and hedgehog signaling. Thus, similar to SMAD1 and SMAD5, which have redundant roles in suppressing granulosa cell tumor development in mice, two type I BMP receptors, BMPR1A and BMPR1B, function together to prevent ovarian tumorigenesis. These studies support a role for a functional BMP signaling axis as a tumor suppressor pathway in the ovary, with BMPR1A and BMPR1B acting downstream of BMP ligands and upstream of BMP receptor SMADs. PMID:20363875

  3. Proliferation of granulosa and thecal cells in germinal disc and non-disc regions during follicular growth in Japanese quail (Coturnix coturnix japonica): bromodeoxyuridine incorporation in situ.

    Science.gov (United States)

    Yoshimura, Y; Okamoto, T; Tamura, T

    1996-05-01

    Proliferation of granulosa and thecal cells was analysed during ovarian follicular growth in laying Japanese quail. The birds were injected intraperitoneally with bromodeoxyuridine (BrdU) 10 or 4 h before ovulation, that is, before or after a preovulatory LH surge, respectively, and incorporation of BrdU by follicular tissues was detected immunocytochemically. Cells labelled with BrdU were seldom seen in the most immature follicles in the ovarian cortex, whereas many granulosa and thecal cells were labelled with BrdU in medium-sized white yolky follicles (approximately 13.3% and 14.4% in granulosa and theca layers, respectively). Ten and four hours before ovulation, the granulosa cells in the germinal disc and non-disc regions of the third largest yellow yolky follicle (F3) were labelled with BrdU (approximately 8.4% and 9.4% in germinal disc; 6.1% and 9.0% in the non-disc region), but only those in the germinal disc region were labelled (approximately 5.4% and 4.0%) in the largest yellow yolky follicle (F1). The percentage of thecal cells labelled with BrdU 4 h before ovulation was significantly higher than the percentage labelled 10 h before ovulation, and was higher in F3 (approximately 11.7%) than in F1 follicles (approximately 5.4%) 4 h before ovulation. These results show that proliferation of granulosa and thecal cells occurs in both germinal disc and non-disc regions in growing follicles, but when a follicle matures proliferation is reduced and in the case of granulosa cells it is restricted to the germinal disc region.

  4. QSAR models for predicting the activity of non-peptide luteinizing hormone-releasing hormone (LHRH) antagonists derived from erythromycin A using quantum chemical properties.

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    Fernández, Michael; Caballero, Julio

    2007-04-01

    Multiple linear regression (MLR) combined with genetic algorithm (GA) and Bayesian-regularized Genetic Neural Networks (BRGNNs) were used to model the binding affinity (pK(I)) of 38 11,12-cyclic carbamate derivatives of 6-O-methylerythromycin A for the Human Luteinizing Hormone-Releasing Hormone (LHRH) receptor using quantum chemical descriptors. A multiparametric MLR equation with good statistical quality was obtained that describes the features relevant for antagonistic activity when the substituent at the position 3 of the erythronolide core was varied. In addition, four-descriptor linear and nonlinear models were established for the whole dataset. Such models showed high statistical quality. However, the BRGNN model was better than the linear model according to the external validation process. In general, our linear and nonlinear models reveal that the binding affinity of the compounds studied for the LHRH receptor is modulated by electron-related terms.

  5. Theca cells and theca-cell conditioned medium inhibit the progression of FSH-induced meiosis of bovine oocytes surrounded by cumulus cells connected to membrana granulosa.

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    van Tol, H T; Bevers, M M

    1998-11-01

    The effect of follicular cells and their conditioned media on the FSH-induced oocyte maturation of oocytes surrounded by cumulus cells connected to the membrana granulosa (COCGs) was investigated. COCGs and cumulus oocyte complexes (COCs) were cultured for 22 hr in M199 supplemented with 0.05 IU FSH/ml in either the presence of pieces of theca cell layer or in the presence of pieces of membrana granulosa. COCGs and COCs were also cultured for 22 hr in either theca-cell conditioned medium (CMt) or in granulosa cell conditioned medium (CMg), both supplemented with 0.05 IU FSH/ml. To investigate the importance of cell-cell contacts between granulosa cells and cumulus cells, oocytes were cultured as COCs in CMt, as COCs in CMt supplemented with pieces of membrana granulosa, or as COCGs in CMt. In all groups the medium was supplemented with 0.05 IU FSH/ml. After culture the nuclear status of the oocytes was assessed using orcein staining. Culture of COCGs in the presence of theca cells as well as in CMt resulted in a significantly decreased proportion of oocytes that had undergone germinal vesicle breakdown (GVBD) at the end of the culture period as compared to the control. Of the oocytes that resumed meiosis in the presence of theca cells or in CMt, the proportion of oocytes that progressed up to the MII stage was significantly reduced. This indicates the production of a meiosis-inhibiting factor by theca cells. Culture with COCs instead of COCGs resulted in comparable results although the effect was less pronounced. The significant effect on the progression of meiosis of oocytes cultured as COCGs or as COCs, obtained in the presence of granulosa cells or in CMg, was much weaker than the effect of theca cells or culture in CMt. Culture of COCs in CMt supplemented with layers of membrana granulosa and 0.05 IU FSH/ml, resulted in significantly less oocytes that resumed meiosis as compared to culture of COCs in CMt. Of the oocytes that showed GVBD, the proportion that

  6. Stimulatory effects of propylthiouracil on pregnenolone production through upregulation of steroidogenic acute regulatory protein expression in rat granulosa cells.

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    Chen, Mei-Chih; Wang, Shyi-Wu; Kan, Shu-Fen; Tsai, Shiow-Chwen; Wu, Yu-Ching; Wang, Paulus S

    2010-12-01

    Propylthiouracil (PTU) is a common and effective clinical medicine for the treatment of hyperthyroidism. Our previous study demonstrated that short-term treatment with PTU inhibits progesterone production in rat granulosa cells. However, our present results indicate that a 16-h treatment with PTU was able to stimulate pregnenolone production in rat granulosa cells, although progesterone production was diminished by PTU through inhibition of 3β-hydroxysteroid dehydrogenase. Notably, we found that PTU treatment enhanced the conversion of cholesterol into pregnenolone, whereas the protein level of the cytochrome P450 side-chain cleavage enzyme (P450scc, which is the enzyme responding to this conversion) was not affected. Interestingly, the levels of steroidogenic acute regulatory protein (StAR) in both total cell lysate and the mitochondrial fraction were significantly increased by PTU treatment. Furthermore, the binding of steroidogenic factor-1 (SF-1) to the StAR promoter region was also enhanced by PTU treatment, which suggests that PTU could upregulate StAR gene expression. In addition to SF-1 regulation, we found that mitogen-activated protein (MAP) kinase kinase activation is an important regulator of PTU-stimulated StAR protein expression, based on the effects of the MEK inhibitor PD98059. In conclusion, these results indicate that PTU plays opposite roles in the production of progesterone and its precursor, pregnenolone. The regulation of negative feedback on speeding the cholesterol transportation and pregnenolone conversion after a 16-h PTU treatment may be the mechanism explaining PTU's inhibition of progesterone production in rat granulosa cells.

  7. The effect of energy balance on the transcriptome of bovine granulosa cells at 60 days postpartum.

    Science.gov (United States)

    Girard, Annie; Dufort, Isabelle; Sirard, Marc-André

    2015-11-01

    Dairy cows expend great amounts of energy during the lactation peak to cope with milk production. A state of negative energy balance (NEB) was suggested as a cause for the suboptimal fertility observed during this period, via an interaction with ovarian function. The objective of this study was to identify the impact of NEB on gene expression in granulosa cells of dairy cows at 60 days postpartum and to suggest a potential treatment to improve ovarian function. Dairy cows at 60 days postpartum from 10 typical medium-sized farms were synchronized using a single injection of prostaglandin. Dominant follicles  were collected 42 hours later by transvaginal aspiration. Blood concentrations of beta-hydroxybutyrate (BHB) on the day of aspiration were used to classify animals into two groups: severe NEB (high BHB, n = 12) and mild NEB (low BHB, n = 12). The transcriptomes of granulosa cells from both groups were contrasted using microarrays, and the differentially expressed genes were analyzed using Ingenuity Pathway Analysis to identify affected functions and potential upstream regulators. Genes linked with cellular organization (KRT4 and PPL), proliferation (TACSTD2), and fatty acids metabolism (VNN2) were downregulated in granulosa cells from animals with severe NEB. Several genes linked to decitabine, a hypomethylating agent, and with beta-estradiol, were downregulated in the severe NEB group. Numerous genes linked to vitamins A and D were also downregulated in this group of cows, suggesting a potential deficiency of these vitamins in dairy cows during the postpartum period. This study supports the idea that energy balance has an impact on follicular dynamics which could be detrimental to resumption of fertility after calving. Copyright © 2015 Elsevier Inc. All rights reserved.

  8. New Report of Two Species of Crabs, Cycloes granulosa and Pugettia vulgaris (Crustacea: Decapoda Collected from Korea

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    Yang, Kea Cheong

    2015-07-01

    Full Text Available Two species of crabs, Cycloes granulosa and Pugettia vulgaris, are described and illustrated for the first time in Korea. The former is the first species of calappoid genus Cycloes and characterized by having a minute lateral spine on the margin of carapace. The latter is a species of majoid crab and similar to P. pellucens. However, it can be distinguished by shorter rostral spines, a smaller hepatic spine, and a carapace entirely covered with short setae. In Korea the calappoid crab now includes seven species of three genera (Calappa, Mursia, and Cycloes and the majoid genus Pugettia consists of six species.

  9. Effects of intramuscular administration of folic acid and vitamin B12 on granulosa cells gene expression in postpartum dairy cows.

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    Gagnon, A; Khan, D R; Sirard, M-A; Girard, C L; Laforest, J-P; Richard, F J

    2015-11-01

    The fertility of dairy cows is challenged during early lactation, and better nutritional strategies need to be developed to address this issue. Combined supplementation of folic acid and vitamin B12 improve energy metabolism in the dairy cow during early lactation. Therefore, the present study was undertaken to explore the effects of this supplement on gene expression in granulosa cells from the dominant follicle during the postpartum period. Multiparous Holstein cows received weekly intramuscular injection of 320 mg of folic acid and 10 mg of vitamin B12 (treated group) beginning 24 (standard deviation=4) d before calving until 56 d after calving, whereas the control group received saline. The urea plasma concentration was significantly decreased during the precalving period, and the concentration of both folate and vitamin B12 were increased in treated animals. Milk production and dry matter intake were not significantly different between the 2 groups. Plasma concentrations of folates and vitamin B12 were increased in treated animals. Daily dry matter intake was not significantly different between the 2 groups before [13.5 kg; standard error (SE)=0.5] and after (23.6 kg; SE=0.9) calving. Average energy-corrected milk tended to be greater in vitamin-treated cows, 39.7 (SE=1.4) and 38.1 (SE=1.3) kg/d for treated and control cows, respectively. After calving, average plasma concentration of β-hydroxybutyrate tended to be lower in cows injected with the vitamin supplement, 0.47 (SE=0.04) versus 0.55 (SE=0.03) for treated and control cows, respectively. The ovarian follicle ≥12 mm in diameter was collected by ovum pick-up after estrus synchronization. Recovered follicular fluid volumes were greater in the vitamin-treated group. A microarray platform was used to investigate the effect of treatment on gene expression of granulosa cells. Lower expression of genes involved in the cell cycle and higher expression of genes associated with granulosa cell differentiation

  10. Effects of orexins A and B on expression of orexin receptors and progesterone release in luteal and granulosa ovarian cells.

    Science.gov (United States)

    Cataldi, Natalia I; Lux-Lantos, Victoria A R; Libertun, Carlos

    2012-10-10

    Orexin-A and orexin-B are neuropeptides controlling sleep-wakefulness, feeding and neuroendocrine functions via their G protein-coupled receptors, orexin-1R and orexin-2R. They are synthesized in the lateral hypothalamus and project throughout the brain. Orexins and orexin receptors have also been described outside the brain. Previously we demonstrated the presence of both receptors in the ovary, their increased expression during proestrous afternoon and the dependence on the gonadotropins. Here we studied the effects of orexins on the mRNA expression of both receptors, by quantitative real-time PCR, on luteal cells from superovulated rat ovaries and granulosa cells from diethylstilbestrol-treated rat ovaries. Effects on progesterone secretion were also measured. In luteal cells, 1 nM of either orexin-A or orexin-B decreased progesterone secretion. Orexin-A treatment increased expression of both orexin-1R and orexin-2R mRNA. The effect on orexin-1R mRNA expression was abolished by an orexin-1R selective receptor antagonist SB-334867 and the effect on orexin-2R mRNA expression was abolished by a selective orexin-2R antagonist JNJ-10397049. Orexin-B did not modify orexin-1R mRNA expression, but increased orexin-2R mRNA expression. The effect of orexin-B on orexin-2R was abolished by a selective orexin-2R antagonist. Neither the expression of orexin receptors nor progesterone secretions by granulosa cells were affected by orexins. FSH, as positive control, increased both steroid hormones secretion, but did not induce the expression of OX receptors in granulosa cells isolated from late preantral/early antral follicles. Finally in ovaries obtained immediately after sacrifice, the expression of orexin-1R and orexin-2R was higher in superovulated rat ovaries compared to control or diethylstilbestrol treated rat ovaries. A selective presence and function of both orexinergic receptors in luteal and granulosa cells is described, suggesting that the orexinergic system may

  11. Torsion theca lutein cyst in association with invasive mole presenting as acute abdomen: a rare case

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    Radhamani S.

    2015-08-01

    Full Text Available Gestational trophoblastic neoplasias (GTN are rare tumours that constitute less than 1% of all gynaecological malignancies. Invasive mole is a distinct subgroup of GTN, which if not diagnosed and treated early, can result in serious complications like uterine perforation and haemoperitoneum. We present a rare case of torsion theca lutein cyst in association with invasive mole of the uterus, which developed following the evacuation of a molar pregnancy with features of continued irregular vaginal bleeding, persistently high betaHcg levels along with acute abdomen. [Int J Reprod Contracept Obstet Gynecol 2015; 4(4.000: 1237-1240

  12. Increases in Plasma Lutein through Supplementation Are Correlated with Increases in Physical Activity and Reductions in Sedentary Time in Older Adults

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    Rebecca L. Thomson

    2014-03-01

    Full Text Available Cross-sectional studies have reported positive relationships between serum lutein concentrations and higher physical activity levels. The purpose of the study was to determine whether increasing plasma lutein levels increases physical activity. Forty-four older adults (BMI, 25.3 ± 2.6 kg/m2; age, 68.8 ± 6.4 year not meeting Australian physical activity guidelines (150 min/week of moderate to vigorous activity were randomized to consume capsules containing 21 mg of lutein or placebo with 250 mL of full-cream milk per day for 4 weeks and encouraged to increase physical activity. Physical activity was assessed by self-report, pedometry and accelerometry (daily activity counts and sedentary time. Exercise self-efficacy was assessed by questionnaire. Thirty-nine participants competed the study (Lutein = 19, Placebo = 20. Lutein increased plasma lutein concentrations compared with placebo (p < 0.001. Absolute and percentage changes in plasma lutein were inversely associated with absolute (r = −0.36, p = 0.03 and percentage changes (r = −0.39, p = 0.02 in sedentary time. Percentage change in plasma lutein was positively associated with the percentage change in average daily activity counts (r = 0.36, p = 0.03. Exercise self-efficacy did not change (p = 0.16. Lutein increased plasma lutein, which was associated with increased physical activity and reduced sedentary time in older adults. Larger trials should evaluate whether Lutein can provide health benefits over the longer term.

  13. The transcriptional targets of mutant FOXL2 in granulosa cell tumours.

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    Roseanne Rosario

    Full Text Available BACKGROUND: Despite their distinct biology, granulosa cell tumours (GCTs are treated the same as other ovarian tumours. Intriguingly, a recurring somatic mutation in the transcription factor Forkhead Box L2 (FOXL2 402C>G has been found in nearly all GCTs examined. This investigation aims to identify the pathogenicity of mutant FOXL2 by studying its altered transcriptional targets. METHODS: The expression of mutant FOXL2 was reduced in the GCT cell line KGN, and wildtype and mutant FOXL2 were overexpressed in the GCT cell line COV434. Total RNA was hybridised to Affymetrix U133 Plus 2 microarrays. Comparisons were made between the transcriptomes of control cells and cells altered by FOXL2 knockdown and overexpression, to detect potential transcriptional targets of mutant FOXL2. RESULTS: The overexpression of wildtype and mutant FOXL2 in COV434, and the silencing of mutant FOXL2 expression in KGN, has shown that mutant FOXL2 is able to differentially regulate the expression of many genes, including two well known FOXL2 targets, StAR and CYP19A. We have shown that many of the genes regulated by mutant FOXL2 are clustered into functional annotations of cell death, proliferation, and tumourigenesis. Furthermore, TGF-β signalling was found to be enriched when using the gene annotation tools GATHER and GeneSetDB. This enrichment was still significant after performing a robust permutation analysis. CONCLUSION: Given that many of the transcriptional targets of mutant FOXL2 are known TGF-β signalling genes, we suggest that deregulation of this key antiproliferative pathway is one way mutant FOXL2 contributes to the pathogenesis of adult-type GCTs. We believe this pathway should be a target for future therapeutic interventions, if outcomes for women with GCTs are to improve.

  14. Prohibitin regulates the FSH signaling pathway in rat granulosa cell differentiation.

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    Chowdhury, Indrajit; Thomas, Kelwyn; Zeleznik, Anthony; Thompson, Winston E

    2016-05-01

    Published results from our laboratory identified prohibitin (PHB), a gene product expressed in granulosa cells (GCs) that progressively increases during follicle maturation. Our current in vitro studies demonstrate that follicle-stimulating hormone (FSH) stimulates Phb expression in rat primary GCs. The FSH-dependent expression of PHB was primarily localized within mitochondria, and positively correlates with the morphological changes in GCs organelles, and synthesis and secretions of estradiol (E2) and progesterone (P4). In order to confirm that PHB plays a regulatory role in rat GC differentiation, endogenous PHB-knockdown studies were carried out in undifferentiated GCs using adenoviral (Ad)-mediated RNA interference methodology. Knockdown of PHB in GCs resulted in the suppression of the key steroidogenic enzymes including steroidogenic acute regulatory protein (StAR), p450 cholesterol side-chain cleavage enzyme (p450scc), 3β-hydroxysteroid dehydrogenase (3β-HSD), and aromatase (Cyp19a1); and decreased E2 and P4 synthesis and secretions in the presence of FSH stimulation. Furthermore, these experimental studies also provided direct evidence that PHB within the mitochondrial fraction in GCs is phosphorylated at residues Y249, T258, and Y259 in response to FSH stimulation. The observed levels of phosphorylation of PHB at Y249, T258, and Y259 were significantly low in GCs in the absence of FSH stimulation. In addition, during GC differentiation FSH-induced expression of phospho-PHB (pPHB) requires the activation of MEK1-ERK1/2 signaling pathway. Taken together, these studies provide new evidence supporting FSH-dependent PHB/pPHB upregulation in GCs is required to sustain the differentiated state of GCs.

  15. PGRMC1 participates in late events of bovine granulosa cells mitosis and oocyte meiosis.

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    Terzaghi, L; Tessaro, I; Raucci, F; Merico, V; Mazzini, G; Garagna, S; Zuccotti, M; Franciosi, F; Lodde, V

    2016-08-02

    Progesterone Receptor Membrane Component 1 (PGRMC1) is expressed in both oocyte and ovarian somatic cells, where it is found in multiple cellular sub-compartments including the mitotic spindle apparatus. PGRMC1 localization in the maturing bovine oocytes mirrors its localization in mitotic cells, suggesting a possible common action in mitosis and meiosis. To test the hypothesis that altering PGRMC1 activity leads to similar defects in mitosis and meiosis, PGRMC1 function was perturbed in cultured bovine granulosa cells (bGC) and maturing oocytes and the effect on mitotic and meiotic progression assessed. RNA interference-mediated PGRMC1 silencing in bGC significantly reduced cell proliferation, with a concomitant increase in the percentage of cells arrested at G2/M phase, which is consistent with an arrested or prolonged M-phase. This observation was confirmed by time-lapse imaging that revealed defects in late karyokinesis. In agreement with a role during late mitotic events, a direct interaction between PGRMC1 and Aurora Kinase B (AURKB) was observed in the central spindle at of dividing cells. Similarly, treatment with the PGRMC1 inhibitor AG205 or PGRMC1 silencing in the oocyte impaired completion of meiosis I. Specifically the ability of the oocyte to extrude the first polar body was significantly impaired while meiotic figures aberration and chromatin scattering within the ooplasm increased. Finally, analysis of PGRMC1 and AURKB localization in AG205-treated oocytes confirmed an altered localization of both proteins when meiotic errors occur. The present findings demonstrate that PGRMC1 participates in late events of both mammalian mitosis and oocyte meiosis, consistent with PGRMC1's localization at the mid-zone and mid-body of the mitotic and meiotic spindle.

  16. Lutein inhibits the function of the transient receptor potential A1 ion channel in different in vitro and in vivo models.

    Science.gov (United States)

    Horváth, Györgyi; Szoke, Éva; Kemény, Ágnes; Bagoly, Teréz; Deli, József; Szente, Lajos; Pál, Szilárd; Sándor, Katalin; Szolcsányi, János; Helyes, Zsuzsanna

    2012-01-01

    Transient receptor potential (TRP) ion channels, such as TRP vanilloid 1 and ankyrin repeat domain 1 (TRPV1 and TRPA1), are expressed on primary sensory neurons. Lutein, a natural tetraterpene carotenoid, can be incorporated into membranes and might modulate TRP channels. Therefore, the effects of the water-soluble randomly methylated-β-cyclodextrin (RAMEB) complex of lutein were investigated on TRPV1 and TRPA1 activation. RAMEB-lutein (100 μM) significantly diminished Ca(2+) influx to cultured rat trigeminal neurons induced by TRPA1 activation with mustard oil, but not by TRPV1 stimulation with capsaicin, as determined with microfluorimetry. Calcitonin gene-related peptide release from afferents of isolated tracheae evoked by mustard oil, but not by capsaicin, was inhibited by RAMEB-lutein. Mustard oil-induced neurogenic mouse ear swelling was also significantly decreased by 100 μg/ml s.c. RAMEB-lutein pretreatment, while capsaicin-evoked edema was not altered. Myeloperoxidase activity indicating non-neurogenic granulocyte accumulation in the ear was not influenced by RAMEB-lutein in either case. It is concluded that lutein inhibits TRPA1, but not TRPV1 stimulation-induced responses on cell bodies and peripheral terminals of sensory neurons in vitro and in vivo. Based on these distinct actions and the carotenoid structure, the ability of lutein to modulate lipid rafts in the membrane around TRP channels can be suggested.

  17. Phototrophic cultivation of a thermo-tolerant Desmodesmus sp. for lutein production: effects of nitrate concentration, light intensity and fed-batch operation.

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    Xie, Youping; Ho, Shih-Hsin; Chen, Ching-Nen Nathan; Chen, Chun-Yen; Ng, I-Son; Jing, Ke-Ju; Chang, Jo-Shu; Lu, Yinghua

    2013-09-01

    Four indigenous thermo-tolerant Desmodesmus sp. strains were examined for their ability to produce lutein. Among them, Desmodesmus sp. F51 was the best strain for this purpose. The medium composition, nitrate concentration and light intensity were manipulated to improve the phototrophic growth and lutein production of Desmodesmus sp. F51. It was found that a nitrogen-sufficient condition was required for lutein accumulation, while a high light intensity enhanced cell growth but caused a decrease in the lutein content. The best cell growth and lutein production occurred when the light intensity and initial nitrate concentration were 600 μmol/m(2)/s and 8.8 mM, respectively. The fed-batch cultivation strategy was shown to further improve lutein production. The highest lutein productivity (3.56±0.10 mg/L/d) and content (5.05±0.20 mg/g) were obtained when pulse-feeding of 2.2 mM nitrate was employed. This study demonstrated the potential of using Desmodesmus sp. F51 as a lutein producer in practical applications.

  18. Lutein and zeaxanthin: Role as macular pigment and factors that control bioavailability from egg yolks and nanoemulsions

    Science.gov (United States)

    Vishwanathan, Rohini

    Lutein and zeaxanthin, two oxygenated carotenoids, exclusively accumulate in the macula, protecting the underlying photoreceptors and retinal pigment epithelial cells from damaging blue radiation of sunlight. As macular pigment, lutein and zeaxanthin are also potent antioxidants protecting the vulnerable regions of retina from free radical injury. Oxidative stress and cumulative light damage play an important role in pathogenesis of age-related macular degeneration (AMD), the leading cause of vision loss in the elderly population. Antioxidant and lutein supplementation has been shown to decrease the risk and prevent the progression of AMD. The egg yolk is a highly bioavailable source of lutein and zeaxanthin and thus a possible contender for AMD prevention and treatment. Consumption of 2 egg yolks/d for 5 weeks was shown herein to significantly increase serum lutein and zeaxanthin concentration and clinically improve macular pigment concentrations at 0.5° retinal eccentricity in an older adult population taking cholesterol-lowering statins. Four egg yolks/d not only raised serum lutein and zeaxanthin significantly but also macular pigment densities at 0.25°, 0.5° and 1° retinal eccentricity. A positive outcome of the 2 egg yolk consumption was the significant increase in serum HDL-C with a tendency of serum LDL-C to decrease, although not significantly. Four egg yolks/d seemed to cross the threshold for dietary cholesterol tolerance as serum LDL-C tended to increase, although not significantly, despite the significant increase in serum HDL-C. There is a strong possibility that greater build up of lutein and zeaxanthin in the macula may have been observed with 2 egg yolks/d if the intervention period was longer than 5 weeks. Addition of up to 2 eggs a day to the diet is suggested to benefit an older adult population, especially those who are already taking cholesterol-lowering statins by (a) building their macular pigment and possibly protect against AMD and (b

  19. Luteinizing hormone and androstendione are independent predictors of ovulation after laparoscopic ovarian drilling: a retrospective cohort study

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    Mayerhofer Klaus

    2009-12-01

    Full Text Available Abstract Background Our objective was to investigate luteinizing hormone, follicle-stimulating hormone, testosterone, and androstenedione as predicitve markers for ovulation after laparoscopic ovarian drilling. Methods We retrospectively analyzed 100 clompihen-resistant patients with the polycystic ovary syndrome who underwent laparoscopic ovarian drilling at our department. The main outcome measure was spontaneous postoperative ovulation within three months after laparoscopic ovarian drilling. In order to predict spontaneous ovulation, we tested the following parameters by use of a univariate followed by a multivariate regression model: Preoperative serum levels of LH, FSH, testosterone, and androstenedione as well as patients' age and body mass index. In addition, we focused on pregnancy and life birth rates. Results Spontaneous ovulation was documented in 71/100 patients (71.0%. In a univariate and multivariate analysis, luteinizing hormone (OR 1.58, 95%CI: 1.30-1.92 and androstenedione (OR 3.03, 95%CI: 1.20-7.67, but not follicle-stimulating hormone and testosterone were independent predictors of ovulation. Using a cut-off for luteinizing hormone and androstenedione of 12.1 IU/l and 3.26 ng/ml, respectively, spontaneous ovulation was observed in 63/70 (90.0% and 36/42 patients (85.7% with elevated and in 8/30 (26.7% and 35/58 (60.3% patients with low luteinizing hormone and androstenedione levels, respectively. The sensitivity, specificity, positive and negatvie predictive values for luteinizing hormone and androstendione as predictors of spontaneous ovulation after ovarian drilling were 88.7% (95%CI: 79.0-95.0%, 75.9% (95%CI: 56.5-89.7%, 90.0% (95%CI: 80.5-95.8%, and 73.3% (95%CI: 54.1-87.7% for luteinizing hormone, and 50.7% (95%CI: 38.6-62.8%, 79.3% (95%CI: 60.3-92.0%, 85.7% (95%CI: 71.5-94.6%, and 39.7% (95%CI: 27.0-53.4% for androstenedione, respectively. Complete one-year follow-up was available for 74/100 patients (74%. We observed a

  20. Lutein epoxide cycle, light harvesting and photoprotection in species of the tropical tree genus Inga.

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    Matsubara, Shizue; Krause, G Heinrich; Seltmann, Martin; Virgo, Aurelio; Kursar, Thomas A; Jahns, Peter; Winter, Klaus

    2008-04-01

    Dynamics and possible function of the lutein epoxide (Lx) cycle, that is, the reversible conversion of Lx to lutein (L) in the light-harvesting antennae, were investigated in leaves of tropical tree species. Photosynthetic pigments were quantified in nine Inga species and species from three other genera. In Inga, Lx levels were high in shade leaves (mostly above 20 mmol mol(-1) chlorophyll) and low in sun leaves. In Virola surinamensis, both sun and shade leaves exhibited very high Lx contents (about 60 mmol mol(-1) chlorophyll). In Inga marginata grown under high irradiance, Lx slowly accumulated within several days upon transfer to deep shade. When shade leaves of I. marginata were briefly exposed to the sunlight, both violaxanthin and Lx were quickly de-epoxidized. Subsequently, overnight recovery occurred only for violaxanthin, not for Lx. In such leaves, containing reduced levels of Lx and increased levels of L, chlorophyll fluorescence induction showed significantly slower reduction of the photosystem II electron acceptor, Q(A), and faster formation as well as a higher level of non-photochemical quenching. The results indicate that slow Lx accumulation in Inga leaves may improve light harvesting under limiting light, while quick de-epoxidation of Lx to L in response to excess light may enhance photoprotection.

  1. Phosphoramide mustard exposure induces DNA adduct formation and the DNA damage repair response in rat ovarian granulosa cells.

    Science.gov (United States)

    Ganesan, Shanthi; Keating, Aileen F

    2015-02-01

    Phosphoramide mustard (PM), the ovotoxic metabolite of the anti-cancer agent cyclophosphamide (CPA), destroys rapidly dividing cells by forming NOR-G-OH, NOR-G and G-NOR-G adducts with DNA, potentially leading to DNA damage. A previous study demonstrated that PM induces ovarian DNA damage in rat ovaries. To investigate whether PM induces DNA adduct formation, DNA damage and induction of the DNA repair response, rat spontaneously immortalized granulosa cells (SIGCs) were treated with vehicle control (1% DMSO) or PM (3 or 6μM) for 24 or 48h. Cell viability was reduced (Padduct was detected after 24h of 6μM PM exposure, while the more cytotoxic G-NOR-G DNA adduct was formed after 48h by exposure to both PM concentrations. Phosphorylated H2AX (γH2AX), a marker of DNA double stranded break occurrence, was also increased by PM exposure, coincident with DNA adduct formation. Additionally, induction of genes (Atm, Parp1, Prkdc, Xrcc6, and Brca1) and proteins (ATM, γH2AX, PARP-1, PRKDC, XRCC6, and BRCA1) involved in DNA repair were observed in both a time- and dose-dependent manner. These data support that PM induces DNA adduct formation in ovarian granulosa cells, induces DNA damage and elicits the ovarian DNA repair response.

  2. Cigarette smoke impairs granulosa cell proliferation and oocyte growth after exposure cessation in young Swiss mice: an experimental study

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    Paixão Larissa LO

    2012-09-01

    Full Text Available Abstract Background Cigarette smoke is associated with decreased female fertility, causing damage to ovarian function and disturbing follicle development. However, the effects of cigarette toxicants on ovarian function depend on duration and intensity of exposure. The aim of this study was to assess the effects of brief, intense exposure to tobacco smoke on granulosa cell number, oocyte growth, and follicle size during puberty in female Swiss mice. Methods Ten female Swiss mice aged 35 days were exposed to tobacco smoke from 3R4F reference research cigarettes. They were exposed to an automatic smoking machine 8 h/day, 7 days/week for 15 days. Ten age-matched controls were kept in a different room and exposed to ambient air. At the end of 15 days, five mice in each group were euthanized and the ovaries were analyzed for follicular morphometry and granulosa cell count. The remaining animals were kept for an additional 30 days for further analysis as an ex-smoker group and control group. Comparison between the two groups was evaluated by the Student’s t-test or a two-way ANOVA followed by Bonferroni post-test was applied for multiple comparisons. Results We found that cigarette smoke impaired antral follicular growth even after exposure cessation (p Conclusions The negative effects of cigarette smoking seem to last even after exposure has been interrupted. Moreover, brief exposure during puberty may induce silent oocyte disruption, which could in turn lead to decreased fecundity rates.

  3. The role of hypoxia and HIF1α in the regulation of STAR-mediated steroidogenesis in granulosa cells.

    Science.gov (United States)

    Kowalewski, Mariusz Pawel; Gram, Aykut; Boos, Alois

    2015-02-05

    The adaptive responses to hypoxia are mediated by hypoxia-inducible factor 1 alpha (HIF1α). Its role, however, in regulating steroidogenesis remains poorly understood. We examined the role of hypoxia and HIF1α in regulating steroid acute regulatory protein (STAR) expression and steroidogenesis in immortalized (KK1) mouse granulosa cells under progressively lowering O2 concentrations (20%, 15%, 10%, 5%, 1%). Basal and dbcAMP-stimulated progesterone synthesis was decreased under severe hypoxia (1% and 5% O2). The partial hypoxia revealed opposing effects, with a significant increase in steroidogenic response at 10% O2 in dbcAMP-treated cells: Star-promoter activity, mRNA and protein expression were increased. The hypoxia-stimulated STAR expression was PKA-dependent. Binding of HIF1α to the Star-promoter was potentiated under partial hypoxia. Inhibition of the transcriptional activity or expression of HIF1α suppressed STAR-expression. HIF1α appears to be a positive regulator of basal and stimulated STAR-expression, which under partial hypoxia is capable of increasing the steroidogenic capacity of granulosa cells.

  4. Mimosine As Well As Serum Starvation Can Be Used for Cell Cycle Synchronization of Sheep Granulosa Cells

    Directory of Open Access Journals (Sweden)

    Fatemeh Sadeghian-Nodoushan

    2014-01-01

    Full Text Available This study was evaluated the effect of different synchronization protocols such as serum starvation for 1–3 days, confluency and chemical inhibitors on synchronization accuracy at G0/G1, apoptosis, and DNA synthesis in sheep granulosa cells. The cells were obtained from ovarian antral follicles of slaughtered sheep and used at first and fifth passages. Flow cytometry analysis showed that confluent cells, serum starvation for 24, 48, and 72 hours, and mimosine treatment significantly increased G0/G1 phase cells when compared to normally growing cells (P<0.05. Nocodazole treatment increased the cell population in the G0/G1 stage when compared with the control group but did not change the G2/M stage population. Treatment of cells with mimosine, nocodazole, and serum starvation in three groups resulted in proliferation arrest (P<0.05. Serum starvation for 72 hours significantly promoted apoptosis in granulosa cells (P<0.05. The results of the primary culture and 5th passage were the same. The use of 48-hour serum starvation and mimosine treatments has been recommended because cell death in these groups was very similar to the control group.

  5. Transfection of isolated rainbow trout, Oncorhynchus mykiss, granulosa cells through chemical transfection and electroporation at 12°C.

    Science.gov (United States)

    Marivin, E; Mourot, B; Loyer, P; Rime, H; Bobe, J; Fostier, A

    2015-09-15

    Over-expression or inhibition of gene expression can be efficiently used to analyse the functions and/or regulation of target genes. Modulation of gene expression can be achieved through transfection of exogenous nucleic acids into target cells. Such techniques require the development of specific protocols to transfect cell cultures with nucleic acids. The aim of this study was to develop a method of transfection suitable for rainbow trout granulosa cells in primary culture. After the isolation of rainbow trout granulosa cells, chemical transfection of cells with a fluorescent morpholino oligonucleotide (MO) was tested using FuGENE HD at 12 °C. Electroporation was also employed to transfect these cells with either a plasmid or MO. Transfection was more efficient using electroporation (with the following settings: 1200 V/40 ms/1p) than chemical transfection, but electroporation by itself was deleterious, resulting in a decrease of the steroidogenic capacity of the cells, measured via estradiol production from its androgenic substrate. The disturbance of cell biology induced by the transfection method per se should be taken into account in data interpretation when investigating the effects of under- or over-expression of candidate genes.

  6. Hedgehog signaling in mouse ovary: Indian hedgehog and desert hedgehog from granulosa cells induce target gene expression in developing theca cells.

    Science.gov (United States)

    Wijgerde, Mark; Ooms, Marja; Hoogerbrugge, Jos W; Grootegoed, J Anton

    2005-08-01

    Follicle development in the mammalian ovary requires interactions among the oocyte, granulosa cells, and theca cells, coordinating gametogenesis and steroidogenesis. Here we show that granulosa cells of growing follicles in mouse ovary act as a source of hedgehog signaling. Expression of Indian hedgehog and desert hedgehog mRNAs initiates in granulosa cells at the primary follicle stage, and we find induced expression of the hedgehog target genes Ptch1 and Gli1, in the surrounding pre-theca cell compartment. Cyclopamine, a highly specific hedgehog signaling antagonist, inhibits this induced expression of target genes in cultured neonatal mouse ovaries. The theca cell compartment remains a target of hedgehog signaling throughout follicle development, showing induced expression of the hedgehog target genes Ptch1, Ptch2, Hip1, and Gli1. In periovulatory follicles, a dynamic synchrony between loss of hedgehog expression and loss of induced target gene expression is observed. Oocytes are unable to respond to hedgehog because they lack expression of the essential signal transducer Smo (smoothened). The present results point to a prominent role of hedgehog signaling in the communication between granulosa cells and developing theca cells.

  7. In vitro production of bovine embryos: cumulus/granulosa cell gene expression patterns point to early atresia as beneficial for oocyte competence

    DEFF Research Database (Denmark)

    Mazzoni, Gianluca; Razza, Eduardo; Pedersen, Hanne S.

    2017-01-01

    In vitro production (IW) of bovine embryos has become widespread technology implemented in cattle breeding and production. Here, we review novel data on cumulus/granulosa cell gene expression, as determined by RNAseq on cellular material from pooled follicular fluids at the single animal level...

  8. Effect of ammonia-generating diet on ovine serum and follicular fluid ammonia and urea levels, serum oestrogen and progesterone concentrations and granulosa cell functions.

    Science.gov (United States)

    Nandi, S; Mondal, S; Pal, D T; Gupta, P S P

    2016-04-01

    This study was undertaken to elucidate the effect of ammonia-generating diet on serum and follicular fluid ammonia and urea levels, serum oestrogen and progesterone concentrations and granulosa cell growth and secretion parameters in ewes (Ovis aries). Ewes were fed with 14% CP diet (control) or ammonia-generating diet or ammonia-generating diet plus soluble sugar. The serum and follicular fluid ammonia and urea level, serum oestrogen and progesterone levels and granulosa cell (obtained from ovaries of slaughtered ewes) growth parameters and secretory activities were estimated. Ammonia-generating diet (high-protein diet) increased the serum ammonia and urea concentration. Supplementation of soluble sugar significantly reduced the ammonia concentration in serum with comparable levels as in control group; however, the urea level in the same group was higher than that observed in control group. Supplementation of soluble sugar significantly reduced the follicular fluid ammonia concentration; however, the level was significantly higher compared to control group. Supplementation of soluble sugar brought down the follicular fluid urea level comparable to that observed in control group. Oestrogen and progesterone levels remained unchanged in ewes fed with different types of diet. Oestrogen and progesterone secretion were significantly lowered from granulosa cells recovered from ewes fed with high ammonia-generating diet. Low metabolic activity and high incidence of apoptosis were observed in granulosa cells obtained from ovaries of ewes fed with ammonia-generating diet.

  9. EVIDENCE FOR EXISTENCE OF IMMUNOGLOBULINS THAT BLOCK OVARIAN GRANULOSA-CELL GROWTH-INVITRO - A PUTATIVE ROLE IN RESISTANT OVARY SYNDROME

    NARCIS (Netherlands)

    VANWEISSENBRUCH, MM; HOEK, A; VAN VLIET BLEEKER, I.; SCHOEMAKER, J; DREXHAGE, H

    1991-01-01

    The sera of 26 patients with premature ovarian failure were examined in order to detect immunoglobulin-G (IgGs) that can block FSH-induced in vitro granulosa cell DNA synthesis via, a Feulgen cytochemical bioassay system. The IgGs of four patients with polycystic ovary-like disease, five postmenopau

  10. Regulation of MicroRNAs, and the Correlations of MicroRNAs and Their Targeted Genes by Zinc Oxide Nanoparticles in Ovarian Granulosa Cells.

    Directory of Open Access Journals (Sweden)

    Yong Zhao

    Full Text Available Zinc oxide (ZnO nanoparticles (NPs have been applied in numerous industrial products and personal care products like sunscreens and cosmetics. The released ZnO NPs from consumer and household products into the environment might pose potential health issues for animals and humans. In this study the expression of microRNAs and the correlations of microRNAs and their targeted genes in ZnO NPs treated chicken ovarian granulosa cells were investigated. ZnSO4 was used as the sole Zn2+ provider to differentiate the effects of NPs from Zn2+. It was found that ZnO-NP-5 μg/ml specifically regulated the expression of microRNAs involved in embryonic development although ZnO-NP-5 μg/ml and ZnSO4-10 μg/ml treatments produced the same intracellular Zn concentrations and resulted in similar cell growth inhibition. And ZnO-NP-5 μg/ml also specifically regulated the correlations of microRNAs and their targeted genes. This is the first investigation that intact NPs in ZnO-NP-5 μg/ml treatment specifically regulated the expression of microRNAs, and the correlations of microRNAs and their targeted genes compared to that by Zn2+. This expands our knowledge for biological effects of ZnO NPs and at the same time it raises the health concerns that ZnO NPs might adversely affect our biological systems, even the reproductive systems through regulation of specific signaling pathways.

  11. Suspected juvenile granulosa cell tumor of scleroring stromal tumor:report of one case and literature review%疑似硬化性间质瘤的幼年型颗粒细胞瘤一例报道并文献复习

    Institute of Scientific and Technical Information of China (English)

    王沛丽; 丁楠; 董驰; 王芳

    2016-01-01

    目的:探讨卵巢幼年型颗粒细胞瘤的临床病理特点。方法:对我院1例疑似硬化性间质瘤的幼年型颗粒细胞瘤行病例报道、病理观察并复习相关文献。结果:患者,女,19岁,下腹痛2月余,B超及MRI检查、肿瘤标志物均提示右侧卵巢良性肿瘤。大体所见,卵巢肿物1个,大小4 cm ×3 cm ×2 cm,切面囊实性,色黄,镜下肿瘤大部分呈实片状、结节状排列,瘤细胞多角形,胞质淡红染,核圆形、卵圆形,深染,偶见核沟,见少数核分裂像,部分区域瘤细胞黄素化较明显,肿瘤组织有滤泡形成,免疫组化病理结果显示免疫表型inhibin-α、CD99、Vimentin、S-100、SMA均呈阳性。术后随访患者12个月,月经恢复正常,无肿瘤复发。结论:卵巢幼年型颗粒细胞瘤是一种罕见的肿瘤,临床工作中应重视其病理特点。%Objective: To discuss clinical and pathological characteristics of the ovarian juvenile granulosa cell tumor. Methods: 1 case in our hospital with suspected juvenile granulosa cell tumor of scleroring stromal tumor was reported and literatures were reviewed.Results: Female, 19-year-old, lower abdominal pain more than 2 months, B ultrasound, MRI and tumor markers all suggested that it was the benign ovarian tumor. An ovarian tumor was found in the right side of ovary and the size is 4 cm × 3 cm × 2 cm. Atfer cutting the surface, it was typically solid with cysts formed. hTe histopathological changes displayed solid lfakes, nodular arrangement. hTe tumor cells were round or oral, cytoplasm stained pink. Nuclear grooves were occasionally conspicuous and mitosis ifgures could be found. Some areas tumor cells were luteinized obvious. hTe immunohistochemical revealed that inhibin-α, CD99, Vimentin, S-100, SMA were all positive. We followed up the patient for 12 months, menstruation was returned to normal and no tumor recurrence.Conclusion: Ovarian juvenile granulosa

  12. Lutein-enriched emulsion-based delivery systems: Influence of pH and temperature on physical and chemical stability.

    Science.gov (United States)

    Davidov-Pardo, Gabriel; Gumus, Cansu Ekin; McClements, David Julian

    2016-04-01

    Lutein may be utilized in foods as a natural pigment or nutraceutical ingredient to improve eye health. Nevertheless, its use is limited by its poor water-solubility and chemical instability. We evaluated the effect of storage temperature and pH on the physical and chemical stability of lutein-enriched emulsions prepared using caseinate. The emulsions (initial droplet diameter=232 nm) remained physically stable at all incubation temperatures (5-70 °C); however the chemical degradation of lutein increased with increasing temperature (activation energy=38 kJ/mol). Solution pH had a major impact on the physical stability of the emulsions, causing droplet aggregation at pH 4 and 5. Conversely, the chemical stability of lutein was largely independent of the pH, with only a slight decrease in degradation at pH 8. This work provides important information for the rational design of emulsion-based delivery systems for a lipophilic natural dye and nutraceutical.

  13. Luteinizing hormone-releasing hormone induces thyroxine release together with testosterone in the neotenic axolotl Ambystoma mexicanum.

    Science.gov (United States)

    Jacobs, G F; Kühn, E R

    1988-09-01

    In male neotenic axolotls Ambystoma mexicanum plasma concentrations of thyroxine (T4) and testosterone were increased following intravenous injection of 10 micrograms luteinizing hormone-releasing hormone. A dose of 50 micrograms influenced only plasma T4 levels. This observation suggests for the first time that a hypothalamic hormone is capable of stimulating the thyroidal axis in the neotenic axolotl.

  14. Depression of plasma luteinizing hormone concentration in quail by the anticholinesterase insecticide parathion

    Science.gov (United States)

    Rattner, B.A.; Clarke, R.N.; Ottinger, M.A.

    1986-01-01

    To examine the effects of parathion on basal plasma luteinizing hormone (LH) concentration, male Japanese quail (Coturnix japonica) were orally intubated with 0, 5 or 10 mg/kg parathion and sacrificed after 4, 8 and 24 hr. At the 5 mg/kg dose, plasma LH levels were reduced at 4 and 8 hr, but returned to control values by 24 hr. Brain acetylcholinesterase activity was substantially reduced by 10 mg/kg parathion (52, 75 and 37% inhibition at 4, 8 and 24 hr, respectively) and plasma LH concentration remained depressed through the 24-hr period. These findings suggest that the organophosphorus insecticide parathion may alter plasma LH concentration in a manner which might impair reproductive activity, and provide indirect evidence for a cholinergic component in the regulation of LH secretion in quail.

  15. Activation of GABA B receptors in the anterior pituitary inhibits prolactin and luteinizing hormone secretion.

    Science.gov (United States)

    Lux-Lantos, V; Rey, E; Libertun, C

    1992-11-01

    Previous work from our laboratory showed that baclofen could lower serum prolactin (PRL) levels acting at the central nervous system. The present experiments were designed to evaluate whether the gamma-aminobutyric acid B agonist was also effective in inhibiting hormone release at the pituitary level. In monolayer cultures of adenohypophyseal dispersed cells, baclofen inhibited basal PRL secretion after 1 or 2 h of incubation. This inhibition was significantly abolished by three antagonists: phaclofen, 3-aminopropyl-phosphonic acid and 4-aminobutylphosphonic acid. Furthermore, baclofen inhibited the thyrotropin-releasing hormone-induced PRL release in a concentration-dependent manner. With regard to gonadotropin secretion, baclofen was unable to modify basal luteinizing hormone (LH) secretion, but significantly inhibited the LH-releasing hormone-induced LH release. These results show that baclofen, in addition to its central neuroendocrine effects, inhibits pituitary hormone secretion, under basal and/or stimulated conditions, by direct action at the pituitary level.

  16. Requirement for specific gravity and creatinine adjustments for urinary steroids and luteinizing hormone concentrations in adolescents.

    Science.gov (United States)

    Singh, Gurmeet K S; Balzer, Ben W R; Desai, Reena; Jimenez, Mark; Steinbeck, Katharine S; Handelsman, David J

    2015-11-01

    Urinary hormone concentrations are often adjusted to correct for hydration status. We aimed to determine whether first morning void urine hormones in growing adolescents require adjustments and, if so, whether urinary creatinine or specific gravity are better adjustments. The study population was adolescents aged 10.1 to 14.3 years initially who provided fasting morning blood samples at 0 and 12 months (n = 343) and first morning urine every three months (n = 644). Unadjusted, creatinine and specific gravity-adjusted hormonal concentrations were compared by Deming regression and Bland-Altman analysis and grouped according to self-rated Tanner stage or chronological age. F-ratios for self-rated Tanner stages and age groups were used to compare unadjusted and adjusted hormonal changes in growing young adolescents. Correlations of paired serum and urinary hormonal concentration of unadjusted and creatinine and specific gravity-adjusted were also compared. Fasting first morning void hormone concentrations correlated well and were unbiased between unadjusted or adjusted by either creatinine or specific gravity. Urine creatinine concentration increases with Tanner stages, age and male gender whereas urine specific gravity was not influenced by Tanner stage, age or gender. Adjustment by creatinine or specific gravity of urinary luteinizing hormone, estradiol, testosterone, dihydrotestosterone and dehydroepiandrosterone concentrations did not improve correlation with paired serum concentrations. Urine steroid and luteinizing hormone concentrations in first morning void samples of adolescents are not significantly influenced by hydration status and may not require adjustments; however, if desired, both creatinine and specific gravity adjustments are equally suitable. © The Author(s) 2015.

  17. Stress and the timing of breeding: glucocorticoid-luteinizing hormones relationships in an arctic seabird.

    Science.gov (United States)

    Goutte, Aurélie; Angelier, Frédéric; Chastel, Céline Clément; Trouvé, Colette; Moe, Børge; Bech, Claus; Gabrielsen, Geir W; Chastel, Olivier

    2010-10-01

    In birds, stressful environmental conditions delay the timing of breeding but the underlying mechanisms are poorly understood. The stress hormone corticosterone appears to be a good candidate for mediating the decision to breed and when to start egg-laying, via a possible inhibition of luteinizing hormone (LH) and sex-steroids production. We used luteinizing hormone releasing hormone (LHRH) challenge in pre-laying male and female Black-legged kittiwakes (Rissa tridactyla) to test whether LH and testosterone secretion were depressed by elevated corticosterone levels. Females bearing high baseline corticosterone levels showed reduced baseline LH levels and a low ability to release LH, following LHRH challenge. Further, females bearing low baseline LH levels and elevated baseline corticosterone levels were more likely to skip breeding. However, non-breeding females were physiologically primed for breeding, since they mounted high LHRH-induced LH release. Egg-laying date was advanced in good body condition females but was unaffected by hormones secretion. In males, corticosterone levels had no effect on LH and/or testosterone secretion and did not affect their decision to breed. Interestingly, males with high LHRH-induced testosterone release bred early. Our study highlights clear sex-differences in the HPG sensitivity to stress hormones in pre-laying kittiwakes. Because females have to store body reserves and to build up the clutch, they would be more sensitive to stress than males. Moreover, intrasexual competition could force male kittiwakes to acquire reproductive readiness earlier in the season than females and to better resist environmental perturbations. We suggest that high testosterone releasing ability would mediate behavioural adjustments such as courtship feeding, which would stimulate early egg-laying in females.

  18. Enhanced Productivity of a Lutein-Enriched Novel Acidophile Microalga Grown on Urea

    Science.gov (United States)

    Casal, Carlos; Cuaresma, Maria; Vega, Jose Maria; Vilchez, Carlos

    2011-01-01

    Coccomyxa acidophila is an extremophile eukaryotic microalga isolated from the Tinto River mining area in Huelva, Spain. Coccomyxa acidophila accumulates relevant amounts of β-carotene and lutein, well-known carotenoids with many biotechnological applications, especially in food and health-related industries. The acidic culture medium (pH < 2.5) that prevents outdoor cultivation from non-desired microorganism growth is one of the main advantages of acidophile microalgae production. Conversely, acidophile microalgae growth rates are usually very low compared to common microalgae growth rates. In this work, we show that mixotrophic cultivation on urea efficiently enhances growth and productivity of an acidophile microalga up to typical values for common microalgae, therefore approaching acidophile algal production towards suitable conditions for feasible outdoor production. Algal productivity and potential for carotenoid accumulation were analyzed as a function of the nitrogen source supplied. Several nitrogen conditions were assayed: nitrogen starvation, nitrate and/or nitrite, ammonia and urea. Among them, urea clearly led to the best cell growth (~4 × 108 cells/mL at the end of log phase). Ammonium led to the maximum chlorophyll and carotenoid content per volume unit (220 μg·mL·1 and 35 μg·mL·1, respectively). Interestingly, no significant differences in growth rates were found in cultures grown on urea as C and N source, with respect to those cultures grown on nitrate and CO2 as nitrogen and carbon sources (control cultures). Lutein accumulated up to 3.55 mg·g·1 in the mixotrophic cultures grown on urea. In addition, algal growth in a shaded culture revealed the first evidence for an active xanthophylls cycle operative in acidophile microalgae. PMID:21339944

  19. Enhanced Productivity of a Lutein-Enriched Novel Acidophile Microalga Grown on Urea

    Directory of Open Access Journals (Sweden)

    Carlos Vilchez

    2010-12-01

    Full Text Available Coccomyxa acidophila is an extremophile eukaryotic microalga isolated from the Tinto River mining area in Huelva, Spain. Coccomyxa acidophila accumulates relevant amounts of b-carotene and lutein, well-known carotenoids with many biotechnological applications, especially in food and health-related industries. The acidic culture medium (pH < 2.5 that prevents outdoor cultivation from non-desired microorganism growth is one of the main advantages of acidophile microalgae production. Conversely, acidophile microalgae growth rates are usually very low compared to common microalgae growth rates. In this work, we show that mixotrophic cultivation on urea efficiently enhances growth and productivity of an acidophile microalga up to typical values for common microalgae, therefore approaching acidophile algal production towards suitable conditions for feasible outdoor production. Algal productivity and potential for carotenoid accumulation were analyzed as a function of the nitrogen source supplied. Several nitrogen conditions were assayed: nitrogen starvation, nitrate and/or nitrite, ammonia and urea. Among them, urea clearly led to the best cell growth (~4 ´ 108 cells/mL at the end of log phase. Ammonium led to the maximum chlorophyll and carotenoid content per volume unit (220 mg·mL-1 and 35 mg·mL-1, respectively. Interestingly, no significant differences in growth rates were found in cultures grown on urea as C and N source, with respect to those cultures grown on nitrate and CO2 as nitrogen and carbon sources (control cultures. Lutein accumulated up to 3.55 mg·g-1 in the mixotrophic cultures grown on urea. In addition, algal growth in a shaded culture revealed the first evidence for an active xanthophylls cycle operative in acidophile microalgae.

  20. Chemical, Lutein and Zeaxanthin Composition of Wheat-yellow Maize Bread for the Prevention of Atherosclerosis

    Directory of Open Access Journals (Sweden)

    Ngozi Innocentia Olaitan

    2015-04-01

    Full Text Available The study examined the effect of substitution of wheat flour with yellow maize flour on the Lutein, Zeaxanthin and proximate composition as well as sensory properties of their bread samples. Standard procedures were used in the production of yellow maize flour. The blend ratios of wheat-yellow maize composite flour used in the production of the bread samples were 100:0 (control, 90:10, 80:20, 70:30, 60:40 and 50:50%, respectively. Results revealed that there were significant (p≤0.05 increases in the lutein 92.4 µg (100:0% to 132.4 µg (50:50% and zeaxanthin 225.0 &mug (100:0% to 580.3 &mug (50:50% content of the bread samples. The proximate composition of the bread samples however, revealed that the carbohydrate, protein and ash content of the bread samples decreased with increase in yellow maize flour. The increase in yellow maize flour however, significantly (p≤0.05 increased the fiber, fat and moisture content of the bread samples. Bread sample B (90:10 was generally the most accepted with an organoleptic score of 8.11, however, sample F (50:50 was the most preferred on the basis of color (7.78, flavor (7.78 and texture (7.58. It would be concluded that substitution of wheat flour with yellow maize flour could be employed to improve on the carotenoids composition of bread, increase their intake and confer protection on consumers against atherosclerosis. The low carbohydrate content of the bread samples could be employed to control spike in blood glucose levels, weight gain and in the management of diabetes. The high fibre content could be harnessed in the prevention and management of obesity and hyperlipideamia.

  1. Identification of cytochrome P-450, and its distribution in the membrana granulosa of the preovulatory follicle, using quantitative cytochemistry.

    Science.gov (United States)

    Zoller, L C; Weisz, J

    1978-07-01

    The distribution of cytochrome P-450 (C-P450), an essential component of the oxidative enzyme system involved in the hydroxylation of steroids, was measured by quantitative cytochemistry in cryostat sections of preovulatory follicles obtained from rats in proestrous. The sections were reacted with medium saturated with carbon monoxide with or without the addition of sodium dithionite. The absorbance spectrum was measured from 400 to 500 nm and a difference spectrum calculated by subtracting the extinction obtained from incubations without sodium dithionite from that obtained in the presence of sodium dithionite. A distinct peak at 450nm was recorded in cells of the peripheral portion of the membrana granulosa (MG) but not in those of the cumulus, providing evidence for the presence and differential distribution of C-P450 in the MG of the preovulatory follicle.

  2. Mioblatoma de células granulosas: Presentación de un caso clínico

    Directory of Open Access Journals (Sweden)

    Nelia Fernández Acosta

    2002-04-01

    Full Text Available El tumor de las células granulosas en una rara entidad; existen numerosas teorías sobre su origen que aún están bajo polémica, aunque la más aceptada es la neurogénica que asimila su procedencia como neuroectodérmico. Se detectó un paciente en consulta con cuadro clínico similar al de esta patología con localización en el dorso de la lengua. La lesión se mostraba submucosa, elevada, no dolorosa y de aspecto áspero. El caso fue remitido a la atención secundaria de cirugía del Instituto Nacional de Oncología y Radiobiología (INOR donde, tras realizar la operación y practicar la biopsia, se corroboró el diagnóstico de un mioblastoma de células granulosas.The tumor of granular cells is a rare entity. There are numerous theories about its origin that are still polemic, though the most accepted is the neurogenic one that assimilates its source as neuroectodermic .A patient with a clinical picture similar to that of this pathology with localization on the dorsum of the tongue was observed at the physician’s office. The lesion was submucous, elevated, unpainful and with a rough aspect. The case was referred to the secondary surgery care service of the National Institute of Oncology and Radiobiology (INOR, in Spanish, where the patient was operated on and a biopsy was performed that confirmed the diagnosis of granular cell myoblastoma.

  3. High salinity tolerance of the Red Sea coral Fungia granulosa under desalination concentrate discharge conditions: an in situ photophysiology experiment

    KAUST Repository

    Van Der Merwe, Riaan

    2014-11-10

    Seawater reverse osmosis desalination concentrate may have chronic and/or acute impacts on the marine ecosystems in the near-field area of the discharge. Environmental impact of the desalination plant discharge is supposedly site- and volumetric- specific, and also depends on the salinity tolerance of the organisms inhabiting the water column in and around a discharge environment. Scientific studies that aim to understand possible impacts of elevated salinity levels are important to assess detrimental effects to organisms, especially for species with no mechanism of osmoregulation, e.g., presumably corals. Previous studies on corals indicate sensitivity toward hypo- and hyper-saline environments with small changes in salinity already affecting coral physiology. In order to evaluate sensitivity of Red Sea corals to increased salinity levels, we conducted a long-term (29 days) in situ salinity tolerance transect study at an offshore seawater reverse osmosis (SWRO) discharge on the coral Fungia granulosa. While we measured a pronounced increase in salinity and temperature at the direct outlet of the discharge structure, effects were indistinguishable from the surrounding environment at a distance of 5 m. Interestingly, corals were not affected by varying salinity levels as indicated by measurements of the photosynthetic efficiency. Similarly, cultured coral symbionts of the genus Symbiodinium displayed remarkable tolerance levels in regard to hypo- and hypersaline treatments. Our data suggest that increased salinity and temperature levels from discharge outlets wear off quickly in the surrounding environment. Furthermore, F. granulosa seem to tolerate levels of salinity that are distinctively higher than reported for other corals previously. It remains to be determined whether Red Sea corals in general display increased salinity tolerance, and whether this is related to prevailing levels of high(er) salinity in the Red Sea in comparison to other oceans.

  4. Phosphoramide mustard exposure induces DNA adduct formation and the DNA damage repair response in rat ovarian granulosa cells

    Energy Technology Data Exchange (ETDEWEB)

    Ganesan, Shanthi, E-mail: shanthig@iastate.edu; Keating, Aileen F., E-mail: akeating@iastate.edu

    2015-02-01

    Phosphoramide mustard (PM), the ovotoxic metabolite of the anti-cancer agent cyclophosphamide (CPA), destroys rapidly dividing cells by forming NOR-G-OH, NOR-G and G-NOR-G adducts with DNA, potentially leading to DNA damage. A previous study demonstrated that PM induces ovarian DNA damage in rat ovaries. To investigate whether PM induces DNA adduct formation, DNA damage and induction of the DNA repair response, rat spontaneously immortalized granulosa cells (SIGCs) were treated with vehicle control (1% DMSO) or PM (3 or 6 μM) for 24 or 48 h. Cell viability was reduced (P < 0.05) after 48 h of exposure to 3 or 6 μM PM. The NOR-G-OH DNA adduct was detected after 24 h of 6 μM PM exposure, while the more cytotoxic G-NOR-G DNA adduct was formed after 48 h by exposure to both PM concentrations. Phosphorylated H2AX (γH2AX), a marker of DNA double stranded break occurrence, was also increased by PM exposure, coincident with DNA adduct formation. Additionally, induction of genes (Atm, Parp1, Prkdc, Xrcc6, and Brca1) and proteins (ATM, γH2AX, PARP-1, PRKDC, XRCC6, and BRCA1) involved in DNA repair were observed in both a time- and dose-dependent manner. These data support that PM induces DNA adduct formation in ovarian granulosa cells, induces DNA damage and elicits the ovarian DNA repair response. - Highlights: • PM forms ovarian DNA adducts. • DNA damage marker γH2AX increased by PM exposure. • PM induces ovarian DNA double strand break repair.

  5. Promotion of glucose utilization by insulin enhances granulosa cell proliferation and developmental competence of porcine oocyte grown in vitro.

    Science.gov (United States)

    Itami, Nobuhiko; Munakata, Yasuhisa; Shirasuna, Koumei; Kuwayama, Takehito; Iwata, Hisataka

    2017-02-01

    In vitro culture of the oocyte granulosa cell complexes (OGCs) from early antral follicles (EAFs) shows granulosa cell (GC) proliferation, but to a lesser extent than that observed in vivo during follicle development. As the number of GCs closely relates to energy sufficiency of the oocytes, enhancement of GC proliferation influences oocyte development. GC proliferation depends on glycolysis and insulin-mediated AKT/mTOR signaling pathway; therefore, addition of culture medium containing insulin and glucose may potentially promote GC proliferation and hence improve oocyte development. In the present study, we assessed the effect of exogenous insulin and glucose concentration on GC proliferation and oocyte energy status as well as developmental abilities of porcine oocytes grown in vitro. In the presence of 5.5 mM of glucose (Low), a comparison of 10 versus 20 μg/ml insulin showed that high insulin enhanced GC proliferation but exhausted glucose from the medium, which resulted in low energy status including lipid and adenosine triphosphate of the oocyte. Whereas, in the presence of 20 μg/ml insulin, medium with 11 mM glucose (High) enhanced GC proliferation and oocyte energy status as well as developmental ability up to the blastocyst stage. Considering that there was no difference in OGCs development observed with medium (10 μg/ml insulin) containing 5.5 versus 11 mM glucose, we concluded that the combination of high insulin and glucose enhanced GC proliferation and energy status of oocytes as well as the developmental ability of the oocytes grown in vitro.

  6. Influence of FSH and hCG on the resumption of meiosis of bovine oocytes surrounded by cumulus cells connected to membrana granulosa.

    Science.gov (United States)

    van Tol, H T; van Eijk, M J; Mummery, C L; van den Hurk, R; Bevers, M M

    1996-10-01

    Cumulus oocyte complexes (COCs) and cumulus oocyte complexes connected to a piece of the membrane granulosa (COCGs) were isolated from bovine antral follicles with a diameter of 2 to 8 mm. After culture of COCGs without gonadotrophic hormones for 22 hr approximately 50% of the oocytes were still in the germinal vesicle (GV) stage. Histology of the COCGs showed that the pieces of the membrana granulosa were free of thecal cells and parts of the basal membrane. This indicates that the membrana granulosa solely inhibits the progression of meiosis. To investigate the effect of gonadotropins on the resumption of meiosis of oocytes from small and medium sized antral follicles, COCs and COCGs were cultured with or without rec-hFSH or hCG. Addition of 0.05 IU rec-hFSH to the culture medium of COCGs resulted in germinal vesicle breakdown in 97.8% of the oocytes compared to 46% in the control group, and an increase of the diameter of the COCs (479 microns vs. 240 microns in the control group). Addition of 0.05 IU hCG to the culture medium had no effect on nuclear maturation (47.2% GV vs. 48.5% GV in the control group) nor on cumulus expansion (246 microns vs. 240 microns in the control group). RT-PCR on cDNA of the follicular wall, cumulus cells, granulosa cells, COCs, and oocytes revealed that mRNA for FSH receptor was present in all cell types except oocytes. mRNA of the LH receptor was detected exclusively in thecal cells. Nucleotide sequence analysis and alignment of the cloned PCR products showed the presence of two isoforms of the FSH receptor mRNA and two isoforms of the LH receptor mRNA. It is concluded that, in vitro, resumption of meiosis of oocytes, originating from small and medium sized antral follicles and meiotically arrested by the membrana granulosa, is triggered by FSH and not by LH. This is supported by the fact that receptors for FSH, but not for LH, are transcribed in the cumulus and granulosa cells of these follicles.

  7. Morphological evidence of apoptosis and the prevalence of apoptotic versus mitotic cells in the membrana granulosa of ovarian follicles during spontaneous and induced atresia in ewes.

    Science.gov (United States)

    Jolly, P D; Smith, P R; Heath, D A; Hudson, N L; Lun, S; Still, L A; Watts, C H; McNatty, K P

    1997-04-01

    Apoptosis is a process by which granulosa cells are thought to be deleted during ovarian follicular atresia. The aims of the present studies, using sheep as the experimental model, were to determine 1) whether morphological changes in cells composing the membrana granulosa during the process of atresia conformed with the general criteria of apoptotic cell death as assessed using tissue sections stained with hematoxylin and eosin; 2) whether cells classified as apoptotic on the basis of their morphology contained fragmented DNA using an in situ 3' end-labeling technique; and 3) the degree of apoptosis and mitosis within the granulosa cell populations of large antral follicles (> or = 3 mm in diameter) during both spontaneous and experimentally induced atresia using stereological methods. The results showed that most degenerate granulosa cells in follicles undergoing atresia display the morphological characteristics of apoptosis, suggesting that this is the most common pathway of cell deletion. Typical features were cells containing nuclei with marginated chromatin; cells with a single small densely staining nucleus (pyknotic appearance); cells with multiple smaller, densely staining nuclear fragments; and densely staining membrane-bound bodies (apoptotic bodies) either singly or in clusters. Cells with morphological features more typical of oncosis or necrosis were sometimes observed, but mainly during the later stages of atresia. All cells classified as apoptotic on the basis of morphological criteria contained fragmented DNA as measured by 3' end-labeling. Apoptotic bodies and/or cells were found in all follicles examined, including those classified as healthy. The overall prevalence of apoptotic cells plus apoptotic bodies expressed as a percentage of the total granulosa cell number per follicle varied from 0.02% to 0.20% in healthy follicles, varied from 0.21% to 2.00% in follicles in early (primary) atresia, and was > 2.0% in follicles in later (secondary

  8. Pattern of human chorionic gonadotropin binding in the polycystic ovary

    Energy Technology Data Exchange (ETDEWEB)

    Brawer, J.; Richard, M.; Farookhi, R. (McGill Univ., Montreal, Quebec (Canada))

    1989-08-01

    The histologic evolution of polycystic ovaries in the estradiol valerate-treated rat coincides with the development of a unique plasma pattern of luteinizing hormone. To assess the role of luteinizing hormone in polycystic ovaries, it is necessary to evaluate the luteinizing hormone sensitivity of the specific tissues in the polycystic ovary. Therefore, we examined the pattern of luteinizing hormone binding sites in polycystic ovaries. Rats at 4 or 8 weeks after estradiol valerate treatment each received an intrajugular injection of iodine 125-labeled human chorionic gonadotropin. Some rats also received a 1000-fold excess of unlabeled human chorionic gonadotropin in the same injection. Ovaries were prepared for autoradiography. Dense accumulations of grains occurred over the theca of normal and atretic secondary follicles in all ovaries and over clusters of secondary interstitial cells. The iodine label was variable over the typically hypertrophied theca of precystic follicles. The theca of definitive cysts showed little or no label. These results indicate that cyst formation coincides with the loss of luteinizing hormone/human chorionic gonadotropin binding to the affected follicles.

  9. Infertility in Female Mice with a Gain-of-Function Mutation in the Luteinizing Hormone Receptor Is Due to Irregular Estrous Cyclicity, Anovulation, Hormonal Alterations, and Polycystic Ovaries

    National Research Council Canada - National Science Library

    Hai, Lan; McGee, Stacey R; Rabideau, Amanda C; Paquet, Marilène; Narayan, Prema

    2015-01-01

    The luteinizing hormone receptor, LHCGR, is essential for fertility in males and females, and genetic mutations in the receptor have been identified that result in developmental and reproductive defects...

  10. Novel C617Y mutation in the 7th transmembrane segment of luteinizing hormone/choriogonadotropin receptor in a Japanese boy with peripheral precocious puberty

    National Research Council Canada - National Science Library

    Nagasaki, Keisuke; Katsumata, Noriyuki; Ogawa, Yohei; Kikuchi, Toru; Uchiyama, Makoto

    2010-01-01

    ... mutations of the LHCGR gene encoding the luteinizing hormone/choriogonadotropin receptor (LH/CGR). The patient is an 8-year-old boy who started to develop pubic hair and penile enlargement at 6 years of age...

  11. Pre-translational regulation of luteinizing hormone receptor in follicular somatic cells of cattle

    Science.gov (United States)

    Marsters, P.; Kendall, N.R.; Campbell, B.K.

    2015-01-01

    Differential regulation of LHR in theca cells (TC) and granulosa cells (GC) is important for normal follicular development. Unlike TC, GC only acquire LH-responsiveness during the later stages of antral follicle development. This study tested the hypothesis that differential LH-responsiveness in these two cell types may be due, in part, to shifts in cellular patterns of alternatively spliced LHR mRNA transcripts which may not be obvious from analysis of total LHR gene expression. It also further explored the role of translation inhibition by an LHR binding protein (LHBP), normally associated with the production of endogenous cholesterol. LHR mRNA variation arises as a result of the alternative splicing of two variable deletion sites (VDS) designated 5′ VDS and 3′ VDS, and it was proposed that differences in cell sensitivity to LH may be due in part to variations in the pattern of the mRNA expression of the receptor variants. The outcomes of the present study support a dynamic multi-facetted regulation of LHR during pre-translation. Not only did the ratio between variants change during antral follicle growth and in vitro cell differentiation but also between TC and GC. Regulation could also be linked to LH concentration feedback mechanisms as the absence of LH caused cultured TC to markedly up-regulate amounts of LHR mRNA. In both TC and GC, LHR mRNA was greatly reduced after treatment to block mevalonate production in the de novo cholesterol pathway, adding further support for a regulatory mechanism linked to enriched cellular amounts of mevalonate kinase. PMID:26507944

  12. HPLC analysis of geometrical isomers of lutein epoxide isolated from dandelion (Taraxacum officinale F. Weber ex Wiggers).

    Science.gov (United States)

    Meléndez-Martínez, Antonio J; Britton, George; Vicario, Isabel M; Heredia, Francisco J

    2006-04-01

    Lutein epoxide has been isolated from petals of dandelion (Taraxacum officinale F. Weber ex Wiggers) by thin-layer chromatography (TLC) on silica to be used for the accurate identification of this carotenoid in other sources. The extract was analyzed by high-performance liquid chromatography (HPLC) using a C(30) column, as a result of which six geometrical isomers were separated. The identification of these isomers was performed on the basis of their UV/vis spectroscopic features in the mobile phase. In quantitative terms, it was observed that all-E-lutein epoxide was the major carotenoid and that there were also high amounts of the (9Z)- and (9'Z)-isomers, although the latter may be an artifact.

  13. ACOMPANHAMENTO CLINICO NUM TUMOR DE CÉLULAS DA GRANULOSA DIAGNOSTICADO EM UMA ÉGUA ATRAVÉS DO ULTRA-SOM

    Directory of Open Access Journals (Sweden)

    Karin Erica Brass

    1994-01-01

    Full Text Available RESUMO Descreve-se o acompanhamento clínico de um caso de tumor de células da granulosa na égua através da ultra-sonografia. Foi constatada uma evolução rápida durante 18 dias para o tumor confirmado, pelo exame macroscópico e histopatológico, após remoção cirúrgica.

  14. Expression of progesterone receptor membrane component-2 within the immature rat ovary and its role in regulating mitosis and apoptosis of spontaneously immortalized granulosa cells.

    Science.gov (United States)

    Griffin, Daniel; Liu, Xiufang; Pru, Cindy; Pru, James K; Peluso, John J

    2014-08-01

    Progesterone receptor membrane component 2 (Pgrmc2) mRNA was detected in the immature rat ovary. By 48 h after eCG, Pgrmc2 mRNA levels decreased by 40% and were maintained at 48 h post-hCG. Immunohistochemical studies detected PGRMC2 in oocytes and ovarian surface epithelial, interstitial, thecal, granulosa, and luteal cells. PGRMC2 was also present in spontaneously immortalized granulosa cells, localizing to the cytoplasm of interphase cells and apparently to the mitotic spindle of cells in metaphase. Interestingly, PGRMC2 levels appeared to decrease during the G1 stage of the cell cycle. Moreover, overexpression of PGRMC2 suppressed entry into the cell cycle, possibly by binding the p58 form of cyclin dependent kinase 11b. Conversely, Pgrmc2 small interfering RNA (siRNA) treatment increased the percentage of cells in G1 and M stage but did not increase the number of cells, which was likely due to an increase in apoptosis. Depleting PGRMC2 did not inhibit cellular (3)H-progesterone binding, but attenuated the ability of progesterone to suppress mitosis and apoptosis. Taken together these studies suggest that PGRMC2 affects granulosa cell mitosis by acting at two specific stages of the cell cycle. First, PGRMC2 regulates the progression from the G0 into the G1 stage of the cell cycle. Second, PGRMC2 appears to localize to the mitotic spindle, where it likely promotes the final stages of mitosis. Finally, siRNA knockdown studies indicate that PGRMC2 is required for progesterone to slow the rate of granulosa cell mitosis and apoptosis. These findings support a role for PGRMC2 in ovarian follicle development.

  15. Toxicity and population structure of the Rough-Skinned Newt (Taricha granulosa) outside the range of an arms race with resistant predators.

    Science.gov (United States)

    Hague, Michael T J; Avila, Leleña A; Hanifin, Charles T; Snedden, W Andrew; Stokes, Amber N; Brodie, Edmund D; Brodie, Edmund D

    2016-05-01

    Species interactions, and their fitness consequences, vary across the geographic range of a coevolutionary relationship. This spatial heterogeneity in reciprocal selection is predicted to generate a geographic mosaic of local adaptation, wherein coevolutionary traits are phenotypically variable from one location to the next. Under this framework, allopatric populations should lack variation in coevolutionary traits due to the absence of reciprocal selection. We examine phenotypic variation in tetrodotoxin (TTX) toxicity of the Rough-Skinned Newt (Taricha granulosa) in regions of allopatry with its TTX-resistant predator, the Common Garter Snake (Thamnophis sirtalis). In sympatry, geographic patterns of phenotypic exaggeration in toxicity and toxin-resistance are closely correlated in prey and predator, implying that reciprocal selection drives phenotypic variation in coevolutionary traits. Therefore, in allopatry with TTX-resistant predators, we expect to find uniformly low levels of newt toxicity. We characterized TTX toxicity in northwestern North America, including the Alaskan panhandle where Ta. granulosa occur in allopatry with Th. sirtalis. First, we used microsatellite markers to estimate population genetic structure and determine if any phenotypic variation in toxicity might be explained by historical divergence. We found northern populations of Ta. granulosa generally lacked population structure in a pattern consistent with northern range expansion after the Pleistocene. Next, we chose a cluster of sites in Alaska, which uniformly lacked genetic divergence, to test for phenotypic divergence in toxicity. As predicted, overall levels of newt toxicity were low; however, we also detected unexpected among- and within-population variation in toxicity. Most notably, a small number of individuals contained large doses of TTX that rival means of toxic populations in sympatry with Th. sirtalis. Phenotypic variation in toxicity, despite limited neutral genetic

  16. Synchronization of ovulation in cyclic gilts with porcine luteinizing hormone (pLH) and its effects on reproductive function.

    Science.gov (United States)

    Degenstein, K L; O'Donoghue, R; Patterson, J L; Beltranena, E; Ambrose, D J; Foxcroft, G R; Dyck, M K

    2008-10-15

    The overall objective was to evaluate the use of porcine luteinizing hormone (pLH) for synchronization of ovulation in cyclic gilts and its effect on reproductive function. In an initial study, four littermate pairs of cyclic gilts were given altrenogest (15 mg/d for 14 d). Gilts received 500 microg cloprostenol (Day 15), 600 IU equine chorionic gonadotropin (eCG) (Day 16) and either 5mg pLH or saline (Control) 80 h after eCG. Blood samples were collected every 4h, from 8h before pLH/saline treatment to the end of estrus. Following estrus detection, transcutaneous real-time ultrasonography and AI, all gilts were slaughtered 6d after the estimated time of ovulation. Peak plasma pLH concentrations (during the LH surge), as well as the amplitude of the LH surge, were greater in pLH-treated gilts than in the control (P=0.01). However, there were no significant differences between treatments in the timing and duration of estrus, or the timing of ovulation within the estrous period. In a second study, 45 cyclic gilts received altrenogest for 14-18d, 600 IU eCG (24h after last altrenogest), and 5mg pLH, 750 IU human chorionic gonadotropin (hCG), or saline, 80 h after eCG. For gilts given pLH or hCG, the diameter of the largest follicle before the onset of ovulation (mean+/-S.E.M.; 8.1+/-0.2 and 8.1+/-0.2mm, respectively) was smaller than in control gilts (8.6+/-0.2mm, P=0.05). The pLH and hCG groups ovulated sooner after treatment compared to the saline-treated group (43.2+/-2.5, 47.6+/-2.5 and 59.5+/-2.5h, respectively; PpLH-treated gilts. Embryo quality (total cell counts and embryo diameter) was not significantly different among groups. In conclusion, pLH reliably synchronized ovulation in cyclic gilts without significantly affecting embryo quality.

  17. Economic Valuation Related to Illegal Extraction of Orchid Cattleya granulosa Within the naTural Park Dom Nivaldo Monte from Natal/RN

    Directory of Open Access Journals (Sweden)

    Adriana Margarida Zanbotto Ramalho

    2010-04-01

    Full Text Available The Atlantic Rain Forest has been the subject of biopiracy and predation of ornamental plants like orchids that reach a high market value. The Natural Park Dom Nivaldo Monte is a fragment of the Atlantic Rain Forest, covering 64 hectares in the city of Natal-RN, which has been the target of this illegal market due to extraction plant of Cattleya granulosa, an endemic orchid endangered. This study aimed to do an economical valuation of the environmental damage caused by the illegal extraction of orchid C. granulosa. The methodology of the survey included the following steps: sample survey of the orchid in the study area and calculation of absolute density and frequency; Economic valuation of environmental damage by using the method proposed by the State Department of Natural Resource Protection – São Paulo – Brazil; proposition the public policies in order to control the environmental impacts in the area. The results pointed out an absolute density and frequency were 1.17 individuals/plot and 8.33%, estimated 1493 individuals, during the period analyzed. The environmental services provided by all individuals of C. granulosa society, in the study area represents a monetary value of R$ 9,555,200.00. It was proposed as a municipal public policies for the preservation of species: increased enforcement in the area, environmental education with the surrounding community, the recovery of damages to the offender, creation of ecological boarder and use of plant biotechnology techniques.

  18. Effect of cortisol on neurophysin I/oxytocin and peptidyl glycine-alpha-amidating mono-oxygenase mRNA expression in bovine luteal and granulosa cells.

    Science.gov (United States)

    Ziolkowska, A; Mlynarczuk, J; Kotwica, J

    2013-01-01

    Cortisol stimulates the synthesis and secretion of oxytocin (OT) from bovine granulosa and luteal cells, but the molecular mechanisms of cortisol action remain unknown. In this study, granulosa cells or luteal cells from days 1-5 and 11-15 of the oestrous cycle were incubated for 4 or 8 h with cortisol (1 x 10(-5), 1 x 10(-7) M). After testing cell viability and hormone secretion (OT, progesterone, estradiol), we studied the effect of cortisol on mRNA expression for precursor of OT (NP-I/OT) and peptidyl glycine-alpha-amidating mono-oxygenase (PGA). The influence of RU 486 (1 x 10(-5) M), a progesterone receptor blocker and inhibitor of the glucocorticosteroid receptor (GR), on the expression for both genes was tested. Cortisol increased the mRNA expression for NP-I/OT and PGA in granulosa cells and stimulated the expression for NP-I/OT mRNA in luteal cells obtained from days 1-5 and days 11-15 of the oestrous cycle. Expression for PGA mRNA was increased only in luteal cells from days 11-15 of the oestrous cycle. In addition, RU 486 blocked the cortisol-stimulated mRNA expression for NP-I/OT and PGA in both types of cells. These data suggest that cortisol affects OT synthesis and secretion in bovine ovarian cells, by acting on the expression of key genes, that may impair ovary

  19. Synthesis and release of luteinizing hormone in vitro: manipulations of Ca2+ environment

    Energy Technology Data Exchange (ETDEWEB)

    Liu, T.C.; Jackson, G.L.

    1985-08-01

    The authors determined if luteinizing hormone (LH) synthesis is Ca2+ dependent and coupled to LH release. They monitored LH synthesis when LH release was stimulated either by specific (gonadotropin-releasing hormone (GnRH)) or nonspecific stimuli (50 mM K+ and 2 or 20 microM Ca2+ ionophore A23187) and inhibited by Ca2+-reduced medium. LH synthesis was estimated by measuring incorporation of (/sup 3/H)glucosamine (glycosylation) and (/sup 14/C)alanine (translation) into total (cell and medium) immunoprecipitable LH by cultured rat anterior pituitary cells. Both GnRH (1 nM) and 50 mM K+ significantly stimulated LH release and glycosylation, but had no effect on LH translation. A23187 also stimulated LH release, but significantly depressed glycosylation of LH and total protein and (/sup 14/C)alanine uptake. Deletion of Ca2+ from the medium depressed both GnRH-induced LH release and glycosylation. Addition of 0.1 mM EGTA to Ca2+-free medium not only inhibited GnRH-induced release and glycosylation of LH but also uptake of precursors and glycosylation and translation of total protein. Thus, glycosylation and release of LH are Ca2+ dependent. Whether parallel changes in LH release and glycosylation reflect a cause and effect relationship remains to be determined.

  20. Biosynthesis and the conjugation of magnetite nanoparticles with luteinizing hormone releasing hormone (LHRH).

    Science.gov (United States)

    Obayemi, J D; Dozie-Nwachukwu, S; Danyuo, Y; Odusanya, O S; Anuku, N; Malatesta, K; Soboyejo, W O

    2015-01-01

    This paper presents the results of an experimental study of the biosynthesis of magnetite nanoparticles (BMNPs) with particle sizes between 10 nm and 60 nm. The biocompatible magnetic nanoparticles are produced from Magnetospirillum magneticum (M.M.) bacteria that respond to magnetic fields. M.M. bacteria were cultured and used to synthesize magnetite nanoparticles. This was done in an enriched magnetic spirillum growth medium (EMSGM) at different pH levels. The nanoparticle concentrations were characterized with UV-Visible (UV-Vis) spectroscopy, while the particle shapes were elucidated via transmission electron microscopy (TEM). The structure of the particles was studied using X-ray diffraction (XRD), while the hydrodynamic radii, particle size distributions and polydispersity of the nanoparticles were characterized using dynamic light scattering (DLS). Carbodiimide reduction was also used to functionalize the BMNPs with a molecular recognition unit (luteinizing hormone releasing hormone, LHRH) that attaches specifically to receptors that are over-expressed on the surfaces of most breast cancer cell types. The resulting nanoparticles were examined using Fourier Transform Infrared (FTIR) spectroscopy and quantitative image analysis. The implications of the results are then discussed for the potential development of magnetic nanoparticles for the specific targeting and treatment of breast cancer.

  1. In vivo pharmacological evaluation of a lactose-conjugated luteinizing hormone releasing hormone analogue.

    Science.gov (United States)

    Moradi, Shayli Varasteh; Varamini, Pegah; Steyn, Frederik; Toth, Istvan

    2015-11-10

    In the current study, the efficacy and pharmacokinetic profile of lactose-conjugated luteinizing hormone releasing hormone (LHRH) was examined following oral administration in male rats. A rapid and sensitive liquid chromatography/mass spectrometry technique was developed and applied for measuring the concentration of lactose[Q(1)][w(6)]LHRH (compound 1) in rat plasma in order to allow measurement of pharmacokinetic parameters. LH release was evaluated using a sandwich ELISA. Maximum serum concentration (Cmax = 0.11 μg/ml) was reached at 2h (Tmax) following oral administration of the compound at 10mg/kg. The half-life was determined to be 2.6h. The absolute bioavailability of the orally administered compound was found to be 14%, which was a remarkable improvement compared to zero-to-low oral bioavailability of the native peptide. Compound 1 was effective in stimulating LH release at 20mg/kg after oral administration. The method was validated at a linear range of 0.01-20.0 μg/ml and a correlation coefficient of r(2) ≥ 0.999. The accuracy and precision values showed the reliability and reproducibility of the method for evaluation of the pharmacokinetic parameters. These findings showed that the lactose derivative of LHRH has a therapeutic potential to be further developed as an orally active therapeutics for the treatment of hormone-dependent diseases.

  2. Effect of supplemental light on growth, prolactin, progesterone and luteinizing hormone in water buffalo ( Bubalus bubalis)

    Science.gov (United States)

    Perera, K. S.; Gwazdauskas, F. C.; Akers, R. M.; McGilliard, M. L.

    1989-06-01

    Fifty non-pregnant Surti buffalo heifers aged between 17 and 42 months ( n=24, 24 months) were randomly assigned to groups subject to either natural daylight +4h supplemental light ( n=25) or natural day light ( n=25), to study changes in growth, serum prolactin (Prl), progesterone (P4) and luteinizing hormone (LH) to supplemental lighting. Ambient temperatures (T) and relative humidity (RH) generally were >27° C and <70% during the day-time, respectively. Light-supplemented heifers had 16.2 kg net body weight (BW) gain at 9 weeks compared to 20.8 kg for controls, but higher mean Prl after 6.5 weeks ( P<0.01), and higher P4 (0.41 vs 0.19 ng/ml; P<0.06) than control heifers. Older heifers had 39.7% greater BW ( P<0.01), but a net 4.3% BW gain compared to a 10.1% gain for younger heifers at 10 weeks. Older, light-supplemented heifers had higher mean P4 (0.63 vs 0.19 ng/ml; P<0.07) than the other groups. These weight and hormonal changes suggest that 4 h supplemental light can alter growth and endocrine function in buffaloes under similar planes of nutrition. While light supplementation did not have a positive effect on body wieght during the 10 week study, body weight and endocrine changes due to supplemental light may be important factors for initiation of reproductive cyclicity.

  3. Quality characteristics and lutein bioavailability from maize and vegetable-based health food.

    Science.gov (United States)

    Ranganathan, Arunkumar; Sheshappa, Mamatha Bangera; Baskaran, Vallikannan

    2014-06-01

    Health food (ready-mix) was prepared from maize and vegetables a source of lutein (L) and zeaxanthin (Z) and studied for its quality characteristics (moisture sorption isotherm, sensory, microbiological, chemical composition, and storage stability) on storage at varying temperatures for 3 months and L+Z bioavailability in mice. Results revealed a decrease in the L+Z level (4.70, 9.24, and 13.85%) of ready-mix stored at 4, 27, and 37°C, respectively. Critical relative humidity and critical moisture content of the product was 64% and 12.24%, respectively. The product is well accepted and was not affected adversely during storage. L+Z bioavailability from ready-mix in mice was higher in plasma (29.4%), liver (58.7%), and eye (14.6%) than control (mice received diet with purified L). To conclude, L+Z in the ready-mix is stable and more bioavailable than control. These findings may help in understanding the importance of simple food processing to improve L bioavailability under its deficient condition among an elderly population.

  4. Cortisol interferes with the estradiol-induced surge of luteinizing hormone in the ewe.

    Science.gov (United States)

    Wagenmaker, Elizabeth R; Breen, Kellie M; Oakley, Amy E; Pierce, Bree N; Tilbrook, Alan J; Turner, Anne I; Karsch, Fred J

    2009-03-01

    Two experiments were conducted to test the hypothesis that cortisol interferes with the positive feedback action of estradiol that induces the luteinizing hormone (LH) surge. Ovariectomized sheep were treated sequentially with progesterone and estradiol to create artificial estrous cycles. Cortisol or vehicle (saline) was infused from 2 h before the estradiol stimulus through the time of the anticipated LH surge in the artificial follicular phase of two successive cycles. The plasma cortisol increment produced by infusion was approximately 1.5 times greater than maximal concentrations seen during infusion of endotoxin, which is a model of immune/inflammatory stress. In experiment 1, half of the ewes received vehicle in the first cycle and cortisol in the second; the others were treated in reverse order. All ewes responded with an LH surge. Cortisol delayed the LH surge and reduced its amplitude, but both effects were observed only in the second cycle. Experiment 2 was modified to provide better control for a cycle effect. Four treatment sequences were tested (cycle 1-cycle 2): vehicle-vehicle, cortisol-cortisol, vehicle-cortisol, cortisol-vehicle. Again, cortisol delayed but did not block the LH surge, and this delay occurred in both cycles. Thus, an elevation in plasma cortisol can interfere with the positive feedback action of estradiol by delaying and attenuating the LH surge.

  5. Cerebral white matter in early puberty is associated with luteinizing hormone concentrations.

    Science.gov (United States)

    Peper, Jiska S; Brouwer, Rachel M; Schnack, Hugo G; van Baal, G Caroline M; van Leeuwen, Marieke; van den Berg, Stéphanie M; Delemarre-Van de Waal, Henriëtte A; Janke, Andrew L; Collins, D Louis; Evans, Alan C; Boomsma, Dorret I; Kahn, René S; Hulshoff Pol, Hilleke E

    2008-08-01

    Puberty is a period in which cerebral white matter grows considerably, whereas gray matter decreases. The first endocrinological marker of puberty in both boys and girls is an increased secretion of luteinizing hormone (LH). Here we investigated the phenotypic association between LH, global and focal gray and white matter in 104 healthy nine-year-old monozygotic and dizygotic twins. Volumetric MRI and voxel-based morphometry were applied to measure global gray and white matter and to estimate relative concentrations of regional cerebral gray and white matter, respectively. A possible common genetic origin of this association (genetic correlation) was examined. Results showed that higher LH levels are associated with a larger global white matter proportion and with higher regional white matter density. Areas of increased white matter density included the cingulum, middle temporal gyrus and splenium of the corpus callosum. No association between LH and global gray matter proportion or regional gray matter density was found. Our data indicate that a common genetic factor underlies the association between LH level and regional white matter density. We suggest that the increase of white matter growth during puberty reported earlier might be directly or indirectly mediated by LH production. In addition, genes involved in LH production may be promising candidate genes in neuropsychiatric illnesses with an onset in early adolescence.

  6. Expression of luteinizing hormone/chorionic gonadotropin receptor in the rat pineal gland.

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    Itoh, Masanori T; Hosaka, Takeshi; Takahashi, Noriyuki; Ishizuka, Bunpei

    2006-08-01

    Luteinizing hormone (LH) influences the secretion of melatonin (N-acetyl-5-methoxytryptamine) from the pineal gland. The present study examined the possible presence of LH/chorionic gonadotropin (CG) receptor in the pineal gland of adult female rats. Reverse transcriptase-polymerase chain reaction analyses demonstrated that LH/CG receptor mRNA is expressed in the pineal gland. Western blotting showed that the pineal gland, like the ovary, contains an 80 kDa receptor protein. Immunohistochemistry revealed that LH/CG receptor, arylalkylamine N-acetyltransferase (a regulatory enzyme in melatonin biosynthesis) and serotonin (a melatonin precursor) are localized primarily to the same cells of the pineal gland. We further found that the levels of pineal LH/CG receptor protein in normal cycling female rats change significantly during the estrous cycle, being lowest at early metestrus. These results demonstrate that LH/CG receptor is expressed in the pineal gland, primarily in melatonin-synthesizing cells, namely pinealocytes. Furthermore, it is suggested that LH influences pineal melatonin secretion through binding to this receptor. In addition, LH/CG receptor levels in the pineal gland are regulated during the estrous cycle under normal physiological conditions.

  7. The problem of anti-doping control of luteinizing hormone in boxing.

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    Llouquet, Jean Louis; Crepin, Nathalie; Lasne, Françoise

    2013-04-01

    Luteinizing hormone (LH) is physiologically produced by the anterior pituitary gland. Male athletes may use pharmaceutical LH for doping since it increases the production of testosterone by testes. This hormone is thus on the World Anti-Doping Agency (WADA) list of substances prohibited for males. Anti-doping laboratories perform the assay of this hormone in urine and report abnormally elevated results. We observed a highly significant prevalence of abnormal results in samples taken after a boxing match. Comparison of the descriptive statistics for 426 LH values observed in boxing and other sports showed significant differences. An experimental study comparing urinary LH levels in 17 boxers before and after a match demonstrated a clear increase after the match. The same observation was made for urinary follicle stimulating hormone (FSH) in all of the eight boxers tested for this other pituitary gonadotropin. These observations have consequences for anti-doping controls, as the reference range for urinary LH levels must take into account the specificities of boxers. They also suggest consequences for the health of boxers. Although to our knowledge such observations have never been described, other pituitary disorders have been reported. Our results deserve further investigation from a medical point of view.

  8. Effect of a complex lutein formula in an animal model for light-induced retinal degeneration.

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    Cheng, Yin-Pin; Ke, Chia-Ying; Kuo, Chih-Chieh; Lee, Yih-Jing

    2016-08-31

    Several retinal degenerative diseases cause vision loss and retinal cell death. Currently, people face prolonged exposure to digital screens, rendering vision protection from light exposure a critical topic. In this study, we designed a complex lutein formula (CLF) by combining several natural compounds: Calendula officinalis, Lycium barbarum, Vaccinium myrtillus, Cassia obtusifolia, and Rhodiola rosea. In addition, we evaluated the protective effects of the formula on retinal functions in an animal model for light-induced retinal degeneration. We employed electroretinography to analyse retinal function, and conducted a histological examination of the morphological changes in the retina treated under various conditions. We revealed that the retinal function in animals exposed to light for 7 days decreased significantly; however, the retinal function of animals that had received the CLF exhibited superior performance, despite light exposure. In addition, a greater portion of the outer nuclear layer (ONL) (i.e. the nuclei of photoreceptors) in these animals was preserved compared with the animals that had not received the formula after 7 days of light exposure. These results revealed that our dietary CLF supplement attenuated retinal function loss resulting from long-term light exposure.

  9. Nitric Oxide and Lutein: Function, Performance, and Protection of Neural Tissue

    Directory of Open Access Journals (Sweden)

    James M. Stringham

    2015-11-01

    Full Text Available The soluble gas neurotransmitter nitric oxide (NO serves many important metabolic and neuroregulatory functions in the retina and brain. Although it is necessary for normal neural function, NO can play a significant role in neurotoxicity. This is often seen in disease states that involve oxidative stress and inflammation of neural tissues, such as age-related macular degeneration and Alzheimer’s disease. The dietary xanthophyll carotenoid lutein (L is a potent antioxidant and anti-inflammatory agent that, if consumed in sufficient amounts, is deposited in neural tissues that require substantial metabolic demand. Some of these specific tissues, such as the central retina and frontal lobes of the brain, are impacted by age-related diseases such as those noted above. The conspicuous correspondence between metabolic demand, NO, and L is suggestive of a homeostatic relationship that serves to facilitate normal function, enhance performance, and protect vulnerable neural tissues. The purpose of this paper is to review the literature on these points.

  10. The expression pattern of microRNAs in granulosa cells of subordinate and dominant follicles during the early luteal phase of the bovine estrous cycle.

    Science.gov (United States)

    Salilew-Wondim, Dessie; Ahmad, Ijaz; Gebremedhn, Samuel; Sahadevan, Sudeep; Hossain, M D Munir; Rings, Franca; Hoelker, Michael; Tholen, Ernst; Neuhoff, Christiane; Looft, Christian; Schellander, Karl; Tesfaye, Dawit

    2014-01-01

    This study aimed to investigate the miRNA expression patterns in granulosa cells of subordinate (SF) and dominant follicle (DF) during the early luteal phase of the bovine estrous cycle. For this, miRNA enriched total RNA isolated from granulosa cells of SF and DF obtained from heifers slaughtered at day 3 and day 7 of the estrous cycle was used for miRNAs deep sequencing. The results revealed that including 17 candidate novel miRNAs, several known miRNAs (n = 291-318) were detected in SF and DF at days 3 and 7 of the estrous cycle of which 244 miRNAs were common to all follicle groups. The let-7 families, bta-miR-10b, bta-miR-26a, bta-miR-99b and bta-miR-27b were among abundantly expressed miRNAs in both SF and DF at both days of the estrous cycle. Further analysis revealed that the expression patterns of 16 miRNAs including bta-miR-449a, bta-miR-449c and bta-miR-222 were differentially expressed between the granulosa cells of SF and DF at day 3 of the estrous cycle. However, at day 7 of the estrous cycle, 108 miRNAs including bta-miR-409a, bta-miR-383 and bta-miR-184 were differentially expressed between the two groups of granulosa cell revealing the presence of distinct miRNA expression profile changes between the two follicular stages at day 7 than day 3 of the estrous cycle. In addition, unlike the SF, marked temporal miRNA expression dynamics was observed in DF groups between day 3 and 7 of the estrous cycle. Target gene prediction and pathway analysis revealed that major signaling associated with follicular development including Wnt signaling, TGF-beta signaling, oocyte meiosis and GnRH signaling were affected by differentially expressed miRNAs. Thus, this study highlights the miRNA expression patterns of granulosa cells in subordinate and dominant follicles that could be associated with follicular recruitment, selection and dominance during the early luteal phase of the bovine estrous cycle.

  11. Inhibition of basal and stimulated progesterone synthesis by dichlorodiphenyldichloroethylene and methoxychlor in a stable pig granulosa cell line.

    Science.gov (United States)

    Crellin, N K; Kang, H G; Swan, C L; Chedrese, P J

    2001-03-01

    The effects of the insecticide dichlorodiphenyldichloroethylene (DDE) and methoxychlor in a stable pig granulosa cell line, JC-410, were investigated. The studies of DDE and methoxychlor were conducted in combination with studies of cholera toxin, the protein kinase A activator that stimulates cAMP and progesterone synthesis and gene expression of P450 cholesterol side chain cleavage (P450scc), which converts cholesterol to pregnenolone. Administration of DDE at 3000 and 10 000 ng ml (-1) was found to decrease progesterone synthesis 0.49- and 0.25-fold, respectively, and to block the stimulatory effect of 100 ng cholera toxin ml (-1), after 24 h incubation. At 1-100 ng ml (-1), methoxychlor did not affect progesterone synthesis after 48 h incubation. However, 1000 ng methoxychlor ml (-1) decreased progesterone synthesis 0.32-fold, and both 100 and 1000 ng methoxychlor ml (-1) blocked the stimulatory effect of cholera toxin. At 3000 and 10 000 ng ml(-1), DDE decreased cAMP synthesis 0.66-and 0.36-fold, respectively. At 300, 3000 and 10 000 ng ml (-1), DDE also decreased cholera toxin-stimulated cAMP synthesis 0.84-, 0.68-, and 0.52-fold, respectively. Administration of 1-100 ng methoxychlor ml (-1) did not affect basal or cholera toxin-stimulated cAMP synthesis. Cholera toxin increased P450scc mRNA 1.4-fold after 24 h incubation, while 3000 and 10 000 ng DDE ml (-1) led to 0.39- and 0.18-fold reductions, respectively. The stimulatory effect of cholera toxin on P450scc mRNA was blocked by 3000 and 10 000 ng DDE ml(-1). Cholera toxin increased P450scc mRNA 3.48-fold after 48 h incubation, while 100 and 1000 ng methoxychlor ml (-1) increased P450scc mRNA 1.79- and 3.0-fold, respectively, and further increased the stimulatory effect of cholera toxin 6.47- and 5.44-fold, respectively. The results of the present study indicate that DDE inhibits granulosa cell steroidogenesis by affecting cAMP production and P450scc gene expression. However, methoxychlor appears to inhibit

  12. Impact of infertility regimens on breast cancer cells: follicle-stimulating hormone and luteinizing hormone lack a direct effect on breast cell proliferation in vitro.

    Science.gov (United States)

    Boukaidi, Samir Alexandre; Cooley, Anne; Hardy, Ashley; Matthews, Laura; Zelivianski, Stanislav; Jeruss, Jacqueline S

    2012-02-01

    To examine the impact of hormones used for controlled ovarian hyperstimulation (COH) on normal and malignant breast cell growth and proliferation. In vitro study of cultured normal and malignant breast cell lines. Academic medical center. None. Normal and malignant breast cell lines cultured in two- and three-dimensional (2D and 3D) systems and treated with follicle-stimulating hormone (FSH), luteinizing hormone (LH), or FSH with LH or human chorionic gonadotropin (hCG). Effects of treatment on cell proliferation in 2D culture using the MTS assay and on colony growth in 3D culture. Compared with untreated cells, normal MCF-10A cells showed a decrease in proliferation and colony size when exposed to a combination of FSH and hCG. The HCC 1937 cells treated with FSH and LH also showed a decrease in colony growth but no change in proliferation. None of the treatments had an effect on the proliferation or colony size of the MCF-7 cells. Follicle-stimulating hormone, LH, and hCG do not appear to cause an increase in cell proliferation or colony growth in either normal or malignant mammary epithelial cell lines. The potential risk for mammary cell transformation associated with these agents may be related to indirect endocrine effects on breast cell physiology. Copyright © 2012 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  13. Faillure in the effect of the analogue (hCG of luteinizing hormone on the luteal angiogenesis in rats (Rattus novergicus

    Directory of Open Access Journals (Sweden)

    Diego Gonçalves Silva

    2013-04-01

    Full Text Available The knowledge of the mechanisms that affect the control of the ovarian activity is essential for the success of reproduction biotechnologies. Although a number of studies have been carried out in which the luteinizing hormone (LH was used to control the ovarian activity, little is known about its influence in the morphology and vascular formation of the corpus luteum, aiming to increase the local blood flow. Thus, the objective of the present experiment was the quantification of the vascular density of corpora lutea (Cls in animals treated with human chorionic gonadotropin (hCG just after ovulation. Therefore, eighteen wistar rats were used in this experiment. Eight rats in the treated group and ten rats in the control group. Corpora lutea were divided into two groups: group (A treated with hCG in the following morning after copulation, and group (B control animals which received an injection of 0.9% sodium chloride solution. Ovaries from each group were used for preparation of histological sections for vascular density qualification. No statistical significance was found between the two groups tested.

  14. Interleukin 1. alpha. inhibits prostaglandin E sub 2 release to suppress pulsatile release of luteinizing hormone but not follicle-stimulating hormone

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    Rettori, V.; McCann, S.M. (Univ. of Texas Southwestern Medical Center, Dallas (United States)); Gimeno, M.F. (CEFYBO, Buenos Aires (Argentina)); Karara, A. (Vanderbilt Univ., Nashville, TN (United States)); Gonzalez, M.C. (Univ. de La Laguno, Tenerife (Spain))

    1991-04-01

    Interleukin 1{alpha} (IL-1{alpha}), a powerful endogenous pyrogen released from monocytes and macrophages by bacterial endotoxin, stimulates corticotropin, prolactin, and somatotropin release and inhibits thyrotropin release by hypothalamic action. The authors injected recombinant human IL-1{alpha} into the third cerebral ventricle, to study its effect on the pulsatile release of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) in conscious, freely moving, ovariectomized rats. Intraventricular injection of 0.25 pmol of IL-1{alpha} caused an almost immediate reduction of plasma LH concentration. To determine the mechanism of the suppression of LH release, mediobasal hypothalamic fragments were incubated in vitro with IL-1{alpha} (10 pM) and the release of LH-releasing hormone (LHRH) and prostaglandin E{sub 2} into the medium was measured by RIA in the presence or absence of nonrepinephrine. 1{alpha} reduced basal LHRH release and blocked LHRH release induced by nonrepinephrine. In conclusion, IL-1{alpha} suppresses LH but not FSH release by an almost complete cessation of pulsatile release of LH in the castrated rat. The mechanism of this effect appears to be by inhibition of prostaglandin E{sub 2}-mediated release of LHRH.

  15. Expression of PUMA in Follicular Granulosa Cells Regulated by FoxO1 Activation During Oxidative Stress.

    Science.gov (United States)

    Liu, Ze-Qun; Shen, Ming; Wu, Wang-Jun; Li, Bo-Jiang; Weng, Qian-Nan; Li, Mei; Liu, Hong-Lin

    2015-06-01

    Many studies have demonstrated that oxidative stress-induced apoptosis is a main cause of follicular atresia. Reactive oxygen species (ROS)-induced granulosa cell (GC) apoptosis is regulated by a variety of signaling pathways involving numerous genes and transcription factors. In this study, we found expression of the p53-upregulated modulator of apoptosis (PUMA), a BH3-only Bcl-2 subfamily protein, in ovarian GCs during oxidative stress. By overexpression and knockdown of Forkhead box O1 (FoxO1), we found that FoxO1 regulates PUMA at the protein level. Moreover, as c-Jun N-terminal kinase (JNK) has been shown to activate FoxO1 by promoting its nuclear import, we used a JNK inhibitor to reduce FoxO1 activation and detected decreased PUMA mes