WorldWideScience

Sample records for luteinized granulosa cells

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

    Directory of Open Access Journals (Sweden)

    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.

  2. Effects of interleukin-8 on estradiol and progesterone production by bovine granulosa cells from large follicles and progesterone production by luteinizing granulosa cells in culture.

    Science.gov (United States)

    Shimizu, Takashi; Kaji, Ayami; Murayama, Chiaki; Magata, Fumie; Shirasuna, Koumei; Wakamiya, Kaori; Okuda, Kiyoshi; Miyamoto, Akio

    2012-01-01

    Interleukin 8 (IL-8) is a chemoattractant involved in the recruitment and activation of neutrophils and is associated with the ovulate process. We examined the possible role of IL-8 in steroid production by bovine granulosa cells before and after ovulation. The concentration of IL-8 in the follicular fluid of estrogen-active dominant (EAD) and pre-ovulatory follicles (POF) was higher than that of small follicles (SF). CXCR1 mRNA expression was higher in the granulosa cells of EAD and POF than that of SF. In contrast, CXCR2 mRNA expression was lower in granulosa cells of EAD and POF than in SF. IL-8 inhibited estradiol (E2) production in follicle-stimulating hormone (FSH)-treated granulosa cells at 48 h of culture. IL-8 also suppressed CYP19A1 mRNA expression in FSH-treated granulosa cells. IL-8 stimulated progesterone (P4) production in luteinizing hormone (LH)-treated granulosa cells at 48 h of culture. Although IL-8 did not alter the expression of genes associated with P4 production, it induced StAR protein expression in LH-treated granulosa cells. The expression of CXCR1 mRNA in corpus luteum (CL) did not change during the luteal phase. In contrast, the expression of CXCR2 mRNA in middle CL was significantly higher than in early and regression CL during the luteal phase. In luteinizing granulosa cells, an in vitro model of granulosa cell luteinization, CXCR2 mRNA expression was downregulated, whereas CXCR1 mRNA expression was unchanged. IL-8 also stimulated P4 production in luteinizing granulosa cells. These data provide evidence that IL-8 functions not only as a chemokine, but also act as a regulator of steroid synthesis in granulosa cells to promote luteinization after ovulation. Copyright © 2011 Elsevier Ltd. All rights reserved.

  3. Effect of Lipopolysaccharide on Progesterone Production during Luteinization of Granulosa and Theca cells In Vitro.

    Science.gov (United States)

    Shimizu, Takashi; Echizenya, Riku; Miyamoto, Akio

    2016-04-01

    The aim of this study is to examine the effect of lipopolysaccharide (LPS) on progesterone production during luteinization of granulosa and theca cells isolated from bovine large follicles. Granulosa and theca cells isolated from large follicles of bovine ovaries were exposed to LPS under appropriate hormone conditions in vitro. Progesterone (P4) production in theca cells, but not granulosa cells, was decreased by long-term exposure of LPS. Long-term exposure of LPS suppressed the gene expression of luteinizing hormone receptor in theca cells. Although long-term exposure of LPS did not affect the expression of steroidogenic acute regulatory protein (StAR) and 3β-hydroxy-steroid dehydrogenase (3β-HSD) genes, it did inhibit the protein expression of StAR and 3β-HSD in theca cells. These findings suggest that theca cells, rather than granulosa cells, are susceptible to LPS during luteinization and that LPS inhibits P4 production by decreasing protein levels of StAR during luteinization of theca cells. © 2016 Wiley Periodicals, Inc.

  4. hCG-dependent regulation of angiogenic factors in human granulosa lutein cells.

    Science.gov (United States)

    Phan, B; Rakenius, A; Pietrowski, D; Bettendorf, H; Keck, C; Herr, D

    2006-07-01

    As prerequisite for development and maintenance of many diseases angiogenesis is of particular interest in medicine. Pathologic angiogenesis takes place in chronic arthritis, collagen diseases, arteriosclerosis, retinopathy associated with diabetes, and particularly in cancers. However, angiogenesis as a physiological process regularly occurs in the ovary. After ovulation the corpus luteum is formed by rapid vascularization of initially avascular granulosa lutein cell tissue. This process is regulated by gonadotropic hormones. In order to gain further insights in the regulatory mechanisms of angiogenesis in the ovary, we investigated these mechanisms in cell culture of human granulosa lutein cells. In particular, we determined the expression and production of several angiogenic factors including tissue inhibitor of matrix metalloproteinases-1 (TIMP-1), Leptin, connective tissue growth factor (CTGF), meningioma-associated complimentary DNA (Mac25), basic fibroblast growth factor (bFGF), and Midkine. In addition, we showed that human chorionic gonadotropin (hCG) has distinct effects on their expression and production. hCG enhances the expression and production of TIMP-1, whereas it downregulates the expression of CTGF and Mac25. Furthermore it decreases the expression of Leptin. Our results provide evidence that hCG determines growth and development of the corpus luteum by mediating angiogenic pathways in human granulosa lutein cells. Hence we describe a further approach to understand the regulation of angiogenesis in the ovary.

  5. Differential regulation of ANG2 and VEGF-A in human granulosa lutein cells by choriogonadotropin.

    Science.gov (United States)

    Pietrowski, D; Keck, C

    2004-04-01

    The growth and development of the corpus luteum after rupture of the follicle is a highly regulated process characterised by a rapid vascularization of the follicle surrounding granulosa cells. Vascularization is regulated by a large number of growth factors and cytokines whereas members of the angiopoietin family and VEGF-A are reported to play a principal role. The gonadotropic hormones luteinizing hormone and choriogonadotropin are reported to be essential for corpus luteum formation. In this study we investigated by RT PCR if the growth factors PGF, PDGF-A, PDGF-B, VEGF-A, VEGF-B, VEGF-C, VEGF-D, ANG1, ANG2, ANG3 and ANG4 are expressed in granulosa cells. We show the expression of VEGF-A, VEGF-B, PDGF-A, ANG1 and ANG2 in granulosa cells. Using RT-PCR and Real-Time PCR we demonstrate that angiopoietin 2 is downregulated in human granulosa cells in vitro after choriogonadotropin treatment whereas the expression of angiopoietin 1 is not significantly altered. The expression of VEGF on the RNA- and on the protein level was determined. It was shown that in granulosa cells VEGF is upregulated after choriogonadotropin treatment on the RNA level and that increasing concentrations of choriogonadotropin from 0 to 10 U/ml leads to an increasing amount of VEGF in the cell culture supernatants. The amount of VEGF in the supernatants reaches a plateau at 0.5 U/ml and is increased only slightly and not significantly after treatment of the cells with 10 U/ml choriogonadotropin compared to 0.5 U/ml. In total these findings suggests that in granulosa cells the mRNA of various growth factors is detectable by RT-PCR and that VEGF-A and ANG2 is regulated by the gonadotropic hormone choriogonadotropin. These findings may add impact on the hypothesis of choriogonadotropin as a novel angiogenic factor.

  6. Hyperglycosylated human chorionic gonadotropin does not increase progesterone production by luteinized granulosa cells.

    Science.gov (United States)

    Crochet, John R; Shah, Anish A; Schomberg, David W; Price, Thomas M

    2012-09-01

    Trophoblast-derived human chorionic gonadotropin (hCG) promotes corpus luteum progesterone (P4) production, and wide ranges of serum P4 levels are noted in various pregnancy outcomes, despite similar hCG concentrations. There are five unique biologically active hCG variants in human pregnancy urine, and previous studies of P4 production in response to hCG have used only preparations containing all isoforms. Understanding exactly which hCG variant is primarily responsible for stimulating corpus luteum steroidogenesis may have great clinical and diagnostic implications, including in the setting of ectopic pregnancy. Our objective was to delineate the role of the standard and hyperglycosylated (H)-hCG isoforms in stimulating P4 production by luteinized granulosa cells. Cell culture, ELISA, and fluorometric-based protein assays were done at Duke University Medical Center. Patients were anonymous oocyte donors. Cultured luteinized granulosa cells were treated with 0.25, 0.5, and 1.0 ng/ml total hCG, which contains all isoforms, purified standard hCG (37.1 kDa), and purified H-hCG (42.8 kDa). P4 produced per total cellular protein (nanograms per microgram) was measured via ELISA and fluorometric protein determination kits. Both total hCG (P = 0.0003) and purified standard hCG (P production. Purified H-hCG did not change the P4 produced per total cellular protein response (P value not significant). Standard hCG stimulated P4 production by cultured granulosa cells and likely supports corpus luteum function via interactions with the LH/hCG receptor. In contrast, H-hCG did not increase P4 production, which indicates a nonsteroidogenic role for this protein during early gestation.

  7. Chorionic gonadotropin regulates the transcript level of VHL, p53, and HIF-2alpha in human granulosa lutein cells.

    Science.gov (United States)

    Herr, D; Keck, C; Tempfer, C; Pietrowski, Detlef

    2004-12-01

    The ovarian corpus luteum plays a critical role in reproduction being the primary source of circulating progesterone. After ovulation the corpus luteum is build by avascular granulosa lutein cells through rapid vascularization regulated by gonadotropic hormones. The present study was performed to investigate whether this process might be influenced by the human chorionic gonadotropin (hCG)-dependent expression of different tumor suppressor genes and hypoxia dependent transcription factors. RNA was isolated from cultured granulosa lutein cells, transcribed into cDNA, and the transcript level of following genes were determined: RB-1, VHL, NF-1, NF-2, Wt-1, p53, APC, and hypoxia inducible factor-1 (HIF-1), -2, and -3alpha. Additionally, the influence of hCG on the expression of VHL, p53, and HIf2alpha were investigated. We demonstrate that in human granulosa lutein cells the tumor suppressor genes RB-1, VHL, NF-1, NF-2, Wt-1, p53, and APC and the hypoxia dependent transcription factors HIF-1alpha, -2alpha, and -3alpha are expressed. In addition, we showed that hCG regulates the expression of p53, VHL, and HIF-2alpha. Our results indicate that hCG may determine the growth and development of the corpus luteum by mediating hypoxic and apoptotic pathways in human granulosa lutein cells. Copyright 2004 Wiley-Liss, Inc.

  8. Small GTPases are involved in sprout formation in human granulosa lutein cells.

    Science.gov (United States)

    Franz, Maximilian B; Daube, Stefanie; Keck, Christoph; Sator, Michael; Pietrowski, Detlef

    2013-04-01

    The corpus luteum (CL), develops from the ruptured follicle after gonadotropin stimulation. Based on intracellular reorganization of the cytoskeleton an human chorionic gonadotropin (hCG) dependent sprouting and migration of luteinizing granulosa cells (LGCs) and endothelial cells is observed. Rho-GTPases are shown to be key regulators of cytoskeletal restructuring. In the present study we analyzed the role of Rho-GTPases in the sprouting activity of LGCs. We used the Rho-GTPase-inhibitors Toxin A and -B and the Cdc42-activator Bradykinin in a LGC-spheroid sprouting assay to determine the effect of these modulators in LGCs. Toxin A and Toxin B reduces sprout formation in LGC spheroids. However, the reduction is less than in hCG treated cells. The usage of Bradykinin demonstrates both, a reduction of sprouts in untreated spheroids and an increase of sprouting in previous hCG treated spheroids. The presented results let us suggest that small Rho-GTPases may regulate the sprouting activity of LGCs after stimulation by hCG and that this mechanism may play a role in CL formation.

  9. Luteinizing hormone-stimulated pituitary adenylate cyclase-activating polypeptide system and its role in progesterone production in human luteinized granulosa cells.

    Science.gov (United States)

    Park, Hyun-Jeong; Choi, Bum-Chae; Song, Sang-Jin; Lee, Dong-Sik; Roh, Jaesook; Chun, Sang-Young

    2010-01-01

    The present study examined the gonadotropin regulation of pituitary adenylate cyclase-activating polypeptide (PACAP) and PACAP type I receptor (PAC(1)-R) expression, and its role in progesterone production in the human luteinized granulosa cells. The stimulation of both PACAP and PAC(1)-R mRNA levels by LH was detected using a competitive reverse transcription-polymerase chain reaction (RT-PCR). PACAP transcript was stimulated by LH reaching maximum levels at 12 hours in a dose dependent manner. LH treatment also stimulated PAC(1)-R mRNA levels within 24 hours. Addition of PACAP-38 (10(-7) M) as well as LH significantly stimulated progesterone production during 48 hours culture. Furthermore, co-treatment with PACAP antagonist partially inhibited LH-stimulated progesterone production. Treatment with vasoactive intestinal peptide, however, did not affect progesterone production. Taken together, the present study demonstrates that LH causes a transient stimulation of PACAP and PAC(1)-R expression and that PACAP stimulates progesterone production in the human luteinized granulosa cells, suggesting a possible role of PACAP as a local ovarian regulator in luteinization.

  10. Dysregulated genes and their functional pathways in luteinized granulosa cells from PCOS patients after cabergoline treatment.

    Science.gov (United States)

    Ferrero, H; Díaz-Gimeno, P; Sebastián-León, P; Faus, A; Gómez, R; Pellicer, A

    2018-04-01

    Polycystic ovarian syndrome (PCOS) is a common reproductive disorder frequently associated with a substantial risk factor for ovarian hyperstimulation syndrome (OHSS). Dopamine receptor 2 (D2) agonists, like cabergoline (Cb2), have been used to reduce the OHSS risk. However, lutein granulosa cells (LGCs) from PCOS patients treated with Cb2 still show a deregulated dopaminergic tone (decreased D2 expression and low dopamine production) and increased vascularization compared to non-PCOS LGCs. Therefore, to understand the PCOS ovarian physiology, it is important to explore the mechanisms that underlie syndrome based on the therapeutic effects of Cb2. Here, LGCs from non-PCOS and PCOS patients were cultured with hCG in the absence/presence of Cb2 ( n  = 12). Subsequently, a transcriptomic-paired design that compared untreated vs treated LGCs within each patient was performed. After transcriptomic analysis, functions and genes were prioritized by systems biology approaches and validated by RT-qPCR. We identified that similar functions were altered in both PCOS and non-PCOS LGCs treated with Cb2; however, PCOS-treated LGCs exhibited more significant changes than non-PCOS. Among the prioritized functions, dopaminergic synapse, vascular endothelial growth factor (VEGF) signaling, apoptosis and ovarian steroidogenesis were highlighted. Finally, network modeling showed CASP9 , VEGFA , AKT1 , CREB , AIF , MAOA , MAPK14 and BMAL1 as key genes implicated in these pathways in Cb2 response, which might be potential biomarkers for further studies in PCOS. © 2018 Society for Reproduction and Fertility.

  11. Luteinized granulosa cell tumor in a seven year old female rottweiler ...

    African Journals Online (AJOL)

    During laparotomy, the kidneys and ovaries were enlarged with metastatic foci to the liver and spleen. Based on the above findings, a confirmatory diagnosis of lutenized granulosa cell tumour was made. Due to evidence of metastasis in several organs, the dog was euthanized with owners consent with intravenous ...

  12. 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, Per Glud; 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. Follicula...

  13. In vitro progesterone production by luteinized human mural granulosa cells is modulated by activation of AMPK and cause of infertility.

    Science.gov (United States)

    Bowdridge, E C; Vernon, M W; Flores, J A; Clemmer, M J

    2017-09-22

    Mural granulosa cells from IVF patients were provided by the West Virginia University Center for Reproductive Medicine in Morgantown, WV. The effect of adenosine monophosphate activated protein kinase (AMPK) activation, primary cause of infertility, age, BMI, and pregnancy outcome on production of progesterone were examined separately. Isolated mural sheets from IVF patients (n = 26) were centrifuged, supernatant discarded, and the pellet re-suspended in 500 μl of DMEM/F12. Mural granulosa cells were plated at 10,000 cells/well in triplicate per treatment group with 300 μl DMEM/F12 media at 37 °C and 5% CO2 in a humidified incubator to permit luteinization. Four days after initial plating, cells were treated with either an AMPK inhibitor, DM; an AMPK activator, AICAR; or hCG. Cells were cultured for 24 h after treatment when medium was collected and frozen at -20 °C until assayed for P4 by radioimmunoassay. The AMPK activator, AICAR, inhibited P4 production (P Progesterone production increased when cells from patients whose primary cause of infertility was a partner having male infertility were treated with hCG compared to control (P = 0.0045), but not in patients with other primary infertility factors (P > 0.05). Additionally, hCG increased P4 production in patients between the ages 30-35 (P = 0.008) and 36-39 (P = 0.04), but not in patients ages 25-29 (P = 0.73). Patients with normal BMI had increased P4 production when treated with hCG (P production from cells of patients who were overweight or obese (P > 0.05). Cells from patients who became pregnant to IVF had greater P4 production when stimulated with hCG than those who did not become pregnant when compared to controls (P > 0.05). Understanding how AMPK activation is regulated in ovarian cells could lead to alternative or novel infertility treatments. Human mural granulosa cells can serve as a valuable model for understanding how AMPK affects P4 production in steroidogenic cells

  14. Luteinizing hormone/chorionic gonadotrophin receptor overexpressed in granulosa cells from polycystic ovary syndrome ovaries is functionally active.

    Science.gov (United States)

    Kanamarlapudi, Venkateswarlu; Gordon, Uma D; López Bernal, Andrés

    2016-06-01

    Polycystic ovarian syndrome (PCOS) is associated with anovulatory infertility. Luteinizing hormone/chorionic gonadotrophin receptor (LHCGR), which is critical for ovulation, has been suggested to be expressed prematurely in the ovarian follicles of women with PCOS. This study aimed to analyse the expression and activity of LHCGR in ovarian granulosa cells from PCOS patients and the involvement of ARF6 small GTPase in LHCGR internalization. Granulosa cells (GC) isolated from follicular fluid collected during oocyte retrieval from normal women (n = 19) and women with PCOS (n = 17) were used to study differences in LHCGR protein expression and activity between normal and PCOS patients. LHCGR expression is up-regulated in GC from PCOS women. LHCGR in PCOS GC is functionally active, as shown by increased cAMP production upon human gonadotrophin (HCG)-stimulation. Moreover, ARF6 is highly expressed in GC from PCOS patients and HCG-stimulation increases the concentrations of active ARF6. The inhibition of ARF6 activation attenuates HCG-induced LHCGR internalization in both normal and PCOS GC, indicating that there are no alterations in LHCGR internalisation in GC from PCOS. In conclusion, the expression and activation of LHCGR and ARF6 are up-regulated in GC from PCOS women but the mechanism of agonist-induced LHCGR internalization is unaltered. Copyright © 2016 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  15. Distinct responses of human granulosa lutein cells after hCG or LH stimulation in a spheroidal cell culture system.

    Science.gov (United States)

    Becker, Julia; Walz, Andrea; Daube, Stefanie; Keck, Christoph; Pietrowski, Detlef

    2007-10-01

    The growth and development of the corpus luteum (CL) is regulated by gonadotropic hormones. It is formed by granulosa cells (GCs), theca cells, and endothelial cells, and is the primary source of circulating progesterone. During early pregnancy only human chorionic gonadotropin (hCG) but not luteinizing hormone (LH) extends the life span of the CL, although hCG and LH interact with the same receptor and have similar actions on the CL. In this study a recently by our group established spheroidal GC culture assay served as a model of CL development on which we compared the actions of the gonadotropic hormones LH and hCG. To find out which signal pathways take part in the hormonal regulation of GC we stimulated GC-spheroids with modulators of protein kinases A and C dependent signaling cascades and determined their impact on sprout forming activity in GC. Our results indicate that PKA-dependent signaling pathways play a major role in mediating the hormonal-induced signaling cascades leading to sprouting in GC. Furthermore, this study strongly indicates that the different effects of hCG and LH in the maintenance of the CL may be reasoned in different signal transduction pathways triggered by hCG or LH. (c) 2007 Wiley-Liss, Inc.

  16. IL-1β Upregulates StAR and Progesterone Production Through the ERK1/2- and p38-Mediated CREB Signaling Pathways in Human Granulosa-Lutein Cells.

    Science.gov (United States)

    Dang, Xuan; Zhu, Qinling; He, Yaqiong; Wang, Yuan; Lu, Yao; Li, Xiaoxue; Qi, Jia; Wu, Hasiximuke; Sun, Yun

    2017-10-01

    The proinflammatory cytokine interleukin-1β (IL-1β) may be involved in several ovulation-associated events, such as protease synthesis, prostaglandin production, and steroidogenesis in granulosa cells. However, the exact effect of IL-1β on progesterone synthesis in granulosa cells and the underlying mechanism remain unclear. By using cultured granulosa-lutein cells collected from women undergoing in vitro fertilization or intracytoplasmic sperm injection, we found that IL-1β upregulated steroidogenic acute regulatory protein (StAR) expression and progesterone synthesis in granulosa-lutein cells, which was comparable with luteinizing hormone effect and could be abolished by an IL-1 receptor antagonist. Moreover, IL-1β activated the phosphorylation of cyclic adenosine monophosphate response element-binding protein (CREB), and knockdown of CREB attenuated the induction of StAR expression and progesterone synthesis by IL-1β in granulosa-lutein cells. Furthermore, IL-1β activated the extracellular signal-regulated kinase (ERK)1/2 and p38 pathways and inhibition of the ERK1/2 and p38 pathways attenuated the IL-1β-induced phosphorylation of CREB, StAR expression, and progesterone synthesis in granulosa-lutein cells. In conclusion, IL-1β could upregulate StAR expression and stimulate progesterone biosynthesis through increase in CREB phosphorylation via activating the ERK1/2 and p38 pathways in human granulosa-lutein cells. Copyright © 2017 Endocrine Society.

  17. BMP6 down-regulates GDNF expression through SMAD1/5 and ERK1/2 signaling pathways in human granulosa-lutein cells.

    Science.gov (United States)

    Zhang, Xin-Yue; Chang, Hsun-Ming; Taylor, Elizabeth L; Leung, Peter C K; Liu, Rui-Zhi

    2018-05-09

    Bone morphogenetic protein 6 (BMP6) is a critical regulator of follicular development that is expressed in mammalian oocytes and granulosa cells. Glial cell line-derived neurotrophic factor (GDNF) is an intraovarian neurotrophic factor that plays an essential role in regulating mammalian oocyte maturation. The aim of this study was to investigate the effect of BMP6 on the regulation of GDNF expression and the potential underlying mechanisms. We used an established immortalized human granulosa cell line (SVOG cells) and primary human granulosa-lutein cells as in vitro cell models. Our results showed that BMP6 significantly down-regulated the expression of GDNF in both SVOG and primary human granulosa-lutein cells. Using dual inhibition approaches (kinase receptor inhibitor and small interfering RNA knockdown), our results showed that both ALK2 and ALK3 are involved in BMP6-induced down-regulation of GDNF. In addition, BMP6 induced the phosphorylation of SMAD1/5/8 and ERK1/2 but not AKT or p38. Among three downstream mediators, both SMAD1 and SMAD5 are involved in BMP6-induced down-regulation of GDNF. Moreover, concomitant knockdown of endogenous SMAD4 and inhibition of ERK1/2 activity completely reversed BMP6-induced down-regulation of GDNF, indicating that both SMAD and ERK1/2 signaling pathways are required for the regulatory effect of BMP6 on GDNF expression. Our findings suggest an additional role for an intrafollicular growth factor in regulating follicular function through their paracrine interactions in human granulosa cells.

  18. Differential responsiveness of luteinized human granulosa cells to gonadotropins and insulin-like growth factor I for induction of aromatase activity

    International Nuclear Information System (INIS)

    Christman, G.M.; Randolph, J.F. Jr.; Peegel, H.; Menon, K.M.

    1991-01-01

    The objective of this study was to examine the in vitro responsiveness of cultured luteinized human granulosa cells over time to insulin-like growth factor 1 (IGF-1), human follicle-stimulating hormone (FSH), and human chorionic gonadotropin (hCG) for the induction of aromatase activity. Granulosa cells were retrieved from preovulatory follicles in patients undergoing in vitro fertilization. Cells were cultured for a period of 72 hours or 10 days. The ability of hCG, human FSH, and/or IGF-I to induce aromatase activity was assayed by the stereospecific release of tritium from [1B-3H]androstenedione. Short-term cultures (72 hours) demonstrated a marked rise in aromatase activity in response to human FSH and IGF-I, whereas a smaller response to hCG was observed. In contrast, 10-day cultures demonstrated responsiveness predominantly to hCG rather than human FSH for the induction of aromatase activity with no remarkable effect of IGF-I. Luteinized human granulosa cells undergo a transformation from an initial human FSH and IGF-I responsive state to an hCG responsive state in long-term cultures

  19. Roles of methyltrienolone (R1881) in AKTs and AR expression patterns of cultured granulosa-lutein cells.

    Science.gov (United States)

    Nekoonam, Saeid; Naji, Mohammad; Mortezaee, Keywan; Amidi, Fardin

    2018-05-11

    AR-mediated androgen signaling plays a key role in female reproductive system. Granulosa-lutein cells (GCs) are the main sites for expression of androgen receptor (AR). There is also a close relation between AKT signaling and AR. Here, we assayed the role for a synthetic AR ligand methyltrienolone (R1881) in expressions of AKTs and AR. Controlled ovarian hyperstimulation (COH) was performed in 20 normal women. Mural GCs were isolated by filtration method, cultured, and passaged. Then, the cells were starved for 48 h with 10% charcoal stripped FBS. The cells were then treated with R1881, bicalutamide (AR blocker), LY294002 (PI3K/AKT pathway blocker), and combination of them for 48 h. Finally, GCs were evaluated for quantitative real-time PCR analysis of AKT1, AKT2, AKT3, and AR, and also Western blot assessment of total AKT and phosphorylated AKT (p-AKT) [Ser473 and Thr308]. Addition of R1881 to the GCs culture showed high expressions of AKT1, AKT2, and AKT3 (P ≤ 0.05 vs LY294002 group and bicalutamide group). Expressions of AKT1 and AKT2 were decreased in the GCs under exposure to bicalutamide or LY294002 (P ≤ 0.05 vs R1881). AKT1, AKT2, and AKT3 showed decreased rates of expressions in the LY294002 + bicalutamide group (P ≤ 0.05 vs R1881). AR, total AKT and p-AKT showed no significant differences between groups. Our findings indicate that 46 h exposure with R1881 could affect AKTs expressions in the GCs of pre-ovulatory phase, but it cannot promote AR expression and AKTs activation. © 2018 Wiley Periodicals, Inc.

  20. Differential insulin and steroidogenic signaling in insulin resistant and non-insulin resistant human luteinized granulosa cells-A study in PCOS patients.

    Science.gov (United States)

    Belani, Muskaan; Deo, Abhilash; Shah, Preeti; Banker, Manish; Singal, Pawan; Gupta, Sarita

    2018-04-01

    Insulin resistance (IR) is one of the significant aberrations in polycystic ovarian syndrome (PCOS), however is only observed in 70%-80% of obese PCOS and 20%-25% of lean PCOS. Hyperinsulinemia accompanies PCOS-IR along with hyperandrogenemia against normal insulin and androgen levels in PCOS-non insulin resistance (NIR). This could possibly be due to defects in the downstream signaling pathways. The study thus aims to unravel insulin and steroidogenic signaling pathways in luteinized granulosa cells isolated from PCOS-IR and NIR vs matched controls. Luteinized granulosa cells from 30 controls and 39 PCOS were classified for IR based on a novel method of down regulation of protein expression of insulin receptor-β (INSR- β) as shown in our previous paper. We evaluated expression of molecules involved in insulin, steroidogenic signaling and lipid metabolism in luteinized granulosa cells followed by analysis of estradiol, progesterone and testosterone in follicular fluid. Protein expression of INSR- β, pIRS (ser 307), PI(3)K, PKC-ζ, pAkt, ERK1/2, pP38MAPK and gene expression of IGF showed differential expression in the two groups. Increased protein expression of PPAR-γ was accompanied by up regulation in SREBP1c, FAS, CPT-1 and ACC-1 genes in PCOS-IR group. Expression of StAR, CYP19A1, 17 β- HSD and 3 β- HSD demonstrated significant decrease along with increase in CYP11A1, FSH-R and LH-R in both the groups. Follicular fluid testosterone increased and progesterone decreased in PCOS-IR group. This study shows how candidate molecules that were differentially expressed, aid in designing targeted therapy against the two phenotypes of PCOS. Copyright © 2018 Elsevier Ltd. All rights reserved.

  1. Expression of antiapoptosis gene survivin in luteinized ovarian granulosa cells of women undergoing IVF or ICSI and embryo transfer: clinical correlations

    Directory of Open Access Journals (Sweden)

    Varras Michail

    2012-09-01

    Full Text Available Abstract Background The purpose of the study was to determine the incidence of survivin gene expression in human granulosa cells during ovarian stimulation in Greek women with normal FSH levels, undergoing IVF or ICSI and to discover any correlation between levels of gene expression and clinical parameters, efficacy of ovulation or outcomes of assisted reproduction. Methods Twenty nine women underwent ovulation induction for IVF or ICSI and ET with standard GnRH analogue-recombinant FSH protocol. Infertility causes were male and tubal factor. Cumulus–mature oocyte complexes were denuded and the granulosa cells were analyzed for each patient separately using quantitative reverse transcription polymerase chain reaction analysis for survivin gene expression with internal standard the ABL gene. Results The ABL and survivin mRNA were detected in granulosa cells in 93.1%. The expression levels of survivin were significantly lower in normal women (male infertility factor compared to women with tubal infertility factor (p = 0.007. There was no additional statistically significant correlation between levels of survivin expression and estradiol levels or dosage of FSH for ovulation induction or number of dominant follicles aspirated or number of retrieved oocytes or embryo grade or clinical pregnancy rates respectively. Conclusions High levels of survivin mRNA expression in luteinized granulosa cells in cases with tubal infertility seem to protect ovaries from follicular apoptosis. A subpopulation of patients with low levels of survivin mRNA in granulosa cells might benefit with ICSI treatment to bypass possible natural barriers of sperm-oocyte interactions.

  2. 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....../l, respectively. VIP at a concentration of 10 nmol/l caused a significant increase in [3H]thymidine incorporation, and at 1000 nmol/l a significant increase in oestradiol secretion was observed. VIP had no effect on progesterone secretion. PHM at the concentrations tested did not influence any of the activities...

  3. Regulation of matrix metalloproteinase-9 (MMP-9), tissue inhibitor of MMP, and progesterone secretion in luteinized granulosa cells from normally ovulating women with polycystic ovary disease.

    Science.gov (United States)

    Ben-Shlomo, Izhar; Goldman, Shlomit; Shalev, Eliezer

    2003-03-01

    To investigate the regulation of MMP-9, TIMP-1, and progesterone via three signal transduction pathways in luteinized granulosa cells from normal ovulatory and PCOD women. In vitro study. Laboratory for Research in Reproductive Sciences, Department of Obstetrics and Gynecology, Ha'Emek Hospital, Afula, Israel. Ten normal ovulatory and 10 women with polycystic ovary disease (PCOD) treated in an assisted reproduction program. Cultured cells were exposed to phorbol 12-myristate 13-acetate (TPA), acting via protein kinase C (PKC), to epidermal growth factor (EGF), acting via protein tyrosine kinase (PTK), and to forskolin, acting via protein kinase A (PKA). Secretion of MMP-9, TIMP-1, and progesterone. Phorbol 12-myristate 13-acetate elicited an increase in MMP-9 and TIMP-1 secretion in both groups and apparently did not affect progesterone secretion. Epidermal growth factor did not change significantly neither MMP-9 nor TIMP-1 secretion but dose dependently decreased MMP-9-TIMP-1 ratio and increased progesterone secretion in the PCOD group. Forskolin inhibited MMP-9 activity and increased TIMP-1 and progesterone secretion in both groups. Progesterone production was inversely related to the ratio of MMP-9-TIMP-1 regardless of cell origin. In this preliminary study, similar and divergent patterns have emerged in the regulation of MMP-9 and TIMP-1 in human luteinized granulosa cells. Repressing MMP-9-TIMP-1 ratio may have an important modulatory effect on progesterone secretion.

  4. Expression of Eph receptor tyrosine kinases and their ligands in human Granulosa lutein cells and human umbilical vein endothelial cells.

    Science.gov (United States)

    Xu, Y; Zagoura, D; Keck, C; Pietrowski, D

    2006-11-01

    Corpus luteum development is regulated by gonadotropins and accompanied by extremely rapid vascularization of the avascular granulosa cell compartiment by endothelial cells (EC). The proliferation of Granulosa cells (GC) and EC is a complex interplay and takes place in a spatially and temporarily coordinated manner. The erythropoietin-producing hepatoma amplified sequence (Eph) receptors and their ligands-the ephrins- are a recently detected family of membrane located protein tyrosine kinases which play a crucial role in the growth and development of nerve and blood vessel network. We report about the mRNA expression pattern of Ephs and their ligands in human GC, in human EC, and in carcinoma cell lines OvCar-3 and Hela. The mRNA of EphA4, EphA7, ephrinA4, ephrinB1 and ephrinB2 was detected in GC and EC, while EphA2 was expressed only in GC. The expression of various Ephs and ephrins did not change in GC after stimulation with human chorion gonadotropin. Our study analyzes for the first time the expression of the complete human Eph/ephriny-system in GC and in EC. The remarkable similarity between these two cell types supports the theory of a functional relationship of EC and GC. In addition, it was shown that hCG is not a major determinant of Eph/ephrin regulation in GC.

  5. Atrazine enhances progesterone production through activation of multiple signaling pathways in FSH-stimulated rat granulosa cells: evidence for premature luteinization.

    Science.gov (United States)

    Pogrmic-Majkic, Kristina; Samardzija, Dragana; Fa, Svetlana; Hrubik, Jelena; Glisic, Branka; Kaisarevic, Sonja; Andric, Nebojsa

    2014-11-01

    Premature luteinization is a possible cause of infertility in women. It is currently unknown whether environmental chemicals can induce changes associated with premature luteinization. Using rat granulosa cells (GC) in vitro, we demonstrated that exposure to atrazine (ATR), a widely used herbicide, causes GC phenotype that resembles that of human premature luteinization. At the end of the 48-h stimulation with FSH, ATR-exposed GC showed (1) higher levels of progesterone, (2) overexpression of luteal markers (Star and Cyp11a1), and (3) an increase in progesterone:estradiol ratio above 1. Mechanistic experiments were conducted to understand the signaling events engaged by ATR that lead to this phenotype. Western blot analysis revealed prolonged phosphorylation of protein kinase B (AKT) and cAMP response element-binding protein (CREB) in ATR- and FSH-exposed GC. An increased level of ERK1/2-dependent transcriptional factor CCATT/enhancer-binding protein beta (CEBPB) was observed after 4 h of ATR exposure. Inhibitors of PI3K (wortmannin) and MEK (U0126) prevented ATR-induced rise in progesterone level and expression of luteal markers in FSH-stimulated GC. Atrazine intensified AKT and CEBPB signaling and caused Star overexpression in forskolin-stimulated GC but not in epidermal growth factor (EGF)-stimulated GC. In the presence of rolipram, a specific inhibitor of phosphodiesterase 4 (PDE4), ATR was not able to further elevate AKT phosphorylation, CEBPB protein level, and Star mRNA in FSH-stimulated GC, suggesting that ATR inhibits PDE4. Overall, this study showed that ATR acts as a FSH sensitizer leading to enhanced cAMP, AKT, and CEBPB signaling and progesterone biosynthesis, which promotes premature luteinization phenotype in GC. © 2014 by the Society for the Study of Reproduction, Inc.

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

    International Nuclear Information System (INIS)

    Shimizu, Takashi; Hirai, Yuko; Murayama, Chiaki; Miyamoto, Akio; Miyazaki, Hitoshi; Miyazaki, Koyomi

    2011-01-01

    Highlights: → Treatment with Per2 and Clock siRNAs decreased the number of granulosa cells and LHr expression. →Per2 siRNA treatment did not stimulate the production of estradiol and expression of P450arom. → 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. → 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.

  7. 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.

  8. PKA- and PKC-dependent regulation of angiopoietin 2 mRNA in human granulosa lutein cells.

    Science.gov (United States)

    Witt, P S; Pietrowski, D; Keck, C

    2004-02-01

    New blood vessels develop from preexisting vessels in response to growth factors or hypoxic conditions. Recent studies have shown that angiopoietin 2 (ANGPT-2) plays an important role in the modulation of angiogenesis and vasculogenesis in humans and mice. The signaling pathways that lead to the regulation of ANGPT-2 are largely unclear. Here, we report that protein kinase C and protein kinase A activators (ADMB, 8-Cl-cAMP) increased the mRNA levels of ANGPT-2 in human Granulosa cells, whereas PKC and PKA Inhibitors (Rp-cAMP, GO 6983) decreased markedly the level of ANGPT-2 mRNA. Due to varying specificity of the modulators for certain protein kinases subunits, we conclude that the conventional PKCs, but not PKC alpha and beta1, the atypical PKCs and the PKA I, are involved in the regulation of ANGPT-2. These findings may help to explain the role of both PKA and PKC dependent signaling cascades in the regulation of ANGPT-2 mRNA.

  9. Growth differentiation factor 9 reverses activin A suppression of steroidogenic acute regulatory protein expression and progesterone production in human granulosa-lutein cells.

    Science.gov (United States)

    Shi, Feng-Tao; Cheung, Anthony P; Klausen, Christian; Huang, He-Feng; Leung, Peter C K

    2010-10-01

    We have reported that growth differentiation factor 9 (GDF9) can enhance activin A (β(A)β(A))-induced inhibin B (αβ(B)) secretion in human granulosa-lutein (hGL) cells, but its effects on steroidogenic acute regulatory protein (StAR), ovarian steroidogenic enzymes, and progesterone production are unknown. We undertook this study to further evaluate GDF9 in this regard. hGL cells from women undergoing in vitro fertilization treatment were cultured with and without small interfering RNA (siRNA) transfection targeted at inhibin α-subunit or GDF9 before treatment with GDF9, activin A, FSH, or combinations. We compared StAR, P450 side-chain cleavage enzyme, and 3β-hydroxysteroid dehydrogenase expression in hGL cells and progesterone levels in culture media after these treatments. mRNA, protein, and hormone levels were assessed with real-time RT-PCR, immunoblotting, and ELISA, respectively. Data were analyzed by ANOVA followed by Tukey's test. Activin A alone reduced basal and FSH-induced progesterone production by decreasing the expression of StAR protein, which regulates the rate-limiting step in steroidogenesis but not P450 side-chain cleavage enzyme and 3β-hydroxysteroid dehydrogenase. GDF9 attenuated these activin A effects on StAR and progesterone. After transfection of α-subunit siRNA, activin A level increased (P progesterone production were attenuated (P progesterone secretion than those observed with activin A treatment alone. GDF9 attenuates the suppressive effects of activin A on StAR expression and progesterone production by increasing the expression of inhibin B, which acts as an activin A competitor.

  10. Growth differentiation factor 3 is induced by bone morphogenetic protein 6 (BMP-6) and BMP-7 and increases luteinizing hormone receptor messenger RNA expression in human granulosa cells.

    Science.gov (United States)

    Shi, Jia; Yoshino, Osamu; Osuga, Yutaka; Akiyama, Ikumi; Harada, Miyuki; Koga, Kaori; Fujimoto, Akihisa; Yano, Tetsu; Taketani, Yuji

    2012-04-01

    To examine the relevance of growth differentiation factor 3 (GDF-3) and bone morphogenetic protein (BMP) cytokines in human ovary. Molecular studies. Research laboratory. Eight women undergoing salpingo-oophorectomy and 30 women undergoing ovarian stimulation for in vitro fertilization. Localizing GDF-3 protein in human ovaries; granulosa cells (GC) cultured with GDF-3, BMP-6, or BMP-7 followed by RNA extraction. The localization of GDF-3 protein in normal human ovaries via immunohistochemical analysis, GDF-3 messenger RNA (mRNA) expression evaluation via quantitative real-time reverse transcription and polymerase chain reaction (RT-PCR), and evaluation of the effect of GDF-3 on leuteinizing hormone (LH) receptor mRNA expression via quantitative real-time RT-PCR. In the ovary, BMP cytokines, of the transforming growth factor beta (TGF-β) superfamily, are known as a luteinization inhibitor by suppressing LH receptor expression in GC. Growth differentiation factor 3, a TGF-β superfamily cytokine, is recognized as an inhibitor of BMP cytokines in other cells. Immunohistochemical analysis showed that GDF-3 was strongly detected in the GC of antral follicles. An in vitro assay revealed that BMP-6 or BMP-7 induced GDF-3 mRNA in GC. Also, GDF-3 increased LH receptor mRNA expression and inhibited the effect of BMP-7, which suppressed the LH receptor mRNA expression in GC. GDF-3, induced with BMP-6 and BMP-7, might play a role in folliculogenesis by inhibiting the effect of BMP cytokines. Copyright © 2012 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  11. BMP4 and BMP7 Suppress StAR and Progesterone Production via ALK3 and SMAD1/5/8-SMAD4 in Human Granulosa-Lutein Cells.

    Science.gov (United States)

    Zhang, Han; Klausen, Christian; Zhu, Hua; Chang, Hsun-Ming; Leung, Peter C K

    2015-11-01

    Adequate production of progesterone by the corpus luteum is critical to the successful establishment of pregnancy. In animal models, bone morphogenetic protein (BMP) 4 and BMP7 have been shown to suppress either basal or gonadotropin-induced progesterone production, depending on the species examined. However, the effects of BMP4 and BMP7 on progesterone production in human granulosa cells are unknown. In the present study, we used immortalized (SVOG) and primary human granulosa-lutein cells to investigate the effects of BMP4 and BMP7 on steroidogenic acute regulatory protein (StAR) expression and progesterone production and to examine the underlying molecular mechanism. Treatment of primary and immortalized human granulosa cells with recombinant BMP4 or BMP7 decreased StAR expression and progesterone accumulation. In SVOG cells, the suppressive effects of BMP4 and BMP7 on StAR expression were blocked by pretreatment with inhibitors of activin receptor-like kinase (ALK)2/3/6 (dorsomorphin) or ALK2/3 (DMH1) but not ALK4/5/7 (SB-431542). Moreover, small interfering RNA-mediated depletion of ALK3, but not ALK2 or ALK6, reversed the effects of BMP4 and BMP7 on StAR expression. Likewise, BMP4- and BMP7-induced phosphorylation of SMAD 1/5/8 was reversed by treatment with DMH1 or small interfering RNA targeting ALK3. Knockdown of SMAD4, the essential common SMAD for BMP/TGF-β signaling, abolished the effects of BMP4 and BMP7 on StAR expression. Our results suggest that BMP4 and BMP7 down-regulate StAR and progesterone production via ALK3 and SMAD1/5/8-SMAD4 signaling in human granulosa-lutein cells.

  12. Expression of miR-15a, miR-145, and miR-182 in granulosa-lutein cells, follicular fluid, and serum of women with polycystic ovary syndrome (PCOS).

    Science.gov (United States)

    Naji, Mohammad; Nekoonam, Saeid; Aleyasin, Ashraf; Arefian, Ehsan; Mahdian, Reza; Azizi, Elham; Shabani Nashtaei, Maryam; Amidi, Fardin

    2018-01-01

    Polycystic ovary syndrome (PCOS) is one of the most common endocrinopathies that affects women in reproductive age. MicroRNAs (miRNAs) play crucial roles in normal function of female reproductive system and folliculogenesis. Deregulated expression of miRNAs in PCOS condition may be significantly implicated in the pathogenesis of PCOS. We determined relative expression of miR-15a, miR-145, and miR-182 in granulosa-lutein cells (GLCs), follicular fluid (FF), and serum of PCOS patients. Human subjects were divided into PCOS (n = 20) and control (n = 21) groups. GLCs, FF, and serum were isolated and stored. RNA isolation was performed and cDNA was reversely transcribed using specific stem-loop RT primers. Relative expression of miRNAs was calculated after normalization against U6 expression. Correlation of miRNAs' expression level with basic clinical features and predictive value of miRNAs in FF and serum were appraised. Relative expression of miR-145 and miR-182 in GLCs was significantly decreased in PCOS, but miR-182 in FF of PCOS patients revealed up-regulated levels. Significant correlations between level of miRNAs in FF and serum and hormonal profile of subjects were observed. MiR-182 in FF showed a significant predictive value with AUC of 0.73, 76.4% sensitivity, and 70.5% specificity which was improved after combination of miR-182 and miR-145. A significant dysregulation of miR-145 and miR-182 in GLCs of PCOS may indicate their involvement in pathogenesis of PCOS. Differential up-regulation of miR-182 in FF of PCOS patients with its promising predictive values for discrimination of PCOS reinforced the importance of studying miRNAs' profile in FF.

  13. (3) Melatonin Protects Oocytes and Granulosa Cells from Reactive Oxygen Species during the Ovulatory Process

    OpenAIRE

    田村, 博史; Hiroshi, TAMURA; 山口大学大学院医学系研究科産科婦人科学; Department of Obstetrics and Gynecology, Yamaguchi University Graduate School of Medicine

    2009-01-01

    Reactive oxygen species (ROS) are produced within the follicle especially during the ovulatory process. ROS play a physiological role in the process of ovulation, e.g. follicle rapture. However, excessive amount of ROS causes oxidative stress and damages oocytes and luteinized granulosa cells. On the other hand, antioxidant defense systems including superoxide dismutase (SOD) or glutathione (GSH) are present in follicles. The balance between ROS and antioxidants within the follicle seems to b...

  14. Juvenile Granulosa Cell Tumour: Anaplastic Variant with Omental Deposits

    Science.gov (United States)

    Rao, Anuradha C.K.; Monappa, Vidya

    2016-01-01

    Juvenile Granulosa Cell Tumour (JGCT) of ovary represents a small fraction of all primary ovarian malignancies. It is a subtype of granulosa cell tumour that is almost always found during the first three decades of life. Histologically, it differs from the typical adult type of granulosa cell tumour. It accounts for 5-15% of all granulosa cell tumours, majority being unilateral. Herein, we describe an unusual histopathological variant of JGCT with numerous large cystic spaces, anaplasia and focal syncytiotrophoblast like giant cells. PMID:27042471

  15. Markers of stem cells in human ovarian granulosa cells: is there a clinical significance in ART?

    Directory of Open Access Journals (Sweden)

    Varras Michail

    2012-11-01

    Full Text Available Abstract Background The purpose of the study was to determine the incidence of gene expression of Oct-4 and DAZL, which are typical markers for stem cells, in human granulosa cells during ovarian stimulation in women with normal FSH levels undergoing IVF or ICSI and to discover any clinical significance of such expression in ART. Methods Twenty one women underwent ovulation induction for IVF or ICSI and ET with standard GnRH analogue-recombinant FSH protocol. Infertility causes were male and tubal factor. Cumulus–mature oocyte complexes were denuded separately and granulosa cells were analyzed for each patient separately using quantitative reverse-transcription–polymerase chain reaction analysis for Oct-4 and DAZL gene expression with G6PD gene as internal standard. Results G6PD and Oct-4 mRNA was detected in the granulosa cells in 47.6% (10/21. The median of Oct-4 mRNA/G6PD mRNA was 1.75 with intra-quarteral range from 0.10 to 98.21. The OCT-4 mRNA expression was statistically significantly correlated with the number of oocytes retrieved; when the Oct-4 mRNA expression was higher, then more than six oocytes were retrieved (p=0.037, Wilcoxon rank-sum. No detection of DAZL mRNA was found in granulosa cells. There was no additional statistically significant correlation between the levels of Oct-4 expression and FSH basal levels or estradiol peak levels or dosage of FSH for ovulation induction. No association was found between the presence or absence of Oct-4 mRNA expression in granulosa cells and ovarian response to gonadotropin stimulation. Also, no influence on pregnancy was observed between the presence or absence of Oct-4 mRNA expression in granulosa cells or to its expression levels accordingly. Conclusions Expression of OCT-4 mRNA, which is a typical stem cell marker and absence of expression of DAZL mRNA, which is a typical germ cell marker, suggest that a subpopulation of luteinized granulosa cells in healthy ovarian follicles (47

  16. Ovarian granulosa cell tumors : histopathology, immunopathology and prognosis

    NARCIS (Netherlands)

    S. Chadha-Ajwani (Savi)

    1987-01-01

    textabstractGranulosa cell tumors (GCT) of the ovary account for 2% of all ovarian tumors. As the name indicates, they are composed of granulosa cells but may also contain an admixture of theca cells. They are potentially malignant but, except for extraovarian spread, which is generally agreed

  17. Knockdown of Progesterone Receptor (PGR) in Macaque Granulosa Cells Disrupts Ovulation and Progesterone Production.

    Science.gov (United States)

    Bishop, Cecily V; Hennebold, Jon D; Kahl, Christoph A; Stouffer, Richard L

    2016-05-01

    Adenoviral vectors (vectors) expressing short-hairpin RNAs complementary to macaque nuclear progesterone (P) receptor PGR mRNA (shPGR) or a nontargeting scrambled control (shScram) were used to determine the role PGR plays in ovulation/luteinization in rhesus monkeys. Nonluteinized granulosa cells collected from monkeys (n = 4) undergoing controlled ovarian stimulation protocols were exposed to either shPGR, shScram, or no virus for 24 h; human chorionic gonadotropin (hCG) was then added to half of the wells to induce luteinization (luteinized granulosa cells [LGCs]; n = 4-6 wells/treatment/monkey). Cells/media were collected 48, 72, and 120 h postvector for evaluation of PGR mRNA and P levels. Addition of hCG increased (P < 0.05) PGR mRNA and medium P levels in controls. However, a time-dependent decline (P < 0.05) in PGR mRNA and P occurred in shPGR vector groups. Injection of shPGR, but not shScram, vector into the preovulatory follicle 20 h before hCG administration during controlled ovulation protocols prevented follicle rupture in five of six monkeys as determined by laparoscopic evaluation, with a trapped oocyte confirmed in three of four follicles of excised ovaries. Injection of shPGR also prevented the rise in serum P levels following the hCG bolus compared to shScram (P < 0.05). Nuclear PGR immunostaining was undetectable in granulosa cells from shPGR-injected follicles, compared to intense staining in shScram controls. Thus, the nuclear PGR appears to mediate P action in the dominant follicle promoting ovulation in primates. In vitro and in vivo effects of PGR knockdown in LGCs also support the hypothesis that P enhances its own synthesis in the primate corpus luteum by promoting luteinization. © 2016 by the Society for the Study of Reproduction, Inc.

  18. Granulosa cell tumor of ovary: US findings

    International Nuclear Information System (INIS)

    Jin, Yong Hyun; Jeon, Hae Jeong; Lee, Chang Dea; Cho, Young Kwon; Kang, Chang Ho; Park, Yong Hyun; Kim, Myung Gyu; Lee, Yeon Hee; Kim, Young Hwa; Lee, Hye Kyung

    1999-01-01

    To describe ultrasonographic findings of ovarian granulosa cell tumor (GCT) and to determine their possible value in the differential diagnosis of ovarian tumors. Sonographic appearances of ten cases of pathologically proven GC Ts were retrospectively reviewed regarding their location, size, outer margin, the echo pattern of the tumor, endometrial thickness, presence of ascites, and metastasis to adjacent tissue or distant sites. 3.0-3.5 MHz trans-abdominal US or 5.0-6.5 MHz transvaginal US were used. The sonographic features could be classified as follows: unilocular cystic mass without nodule or septation (type 1), multilocular cystic mass (type 2), and solid mass (type 3). Pathologically nine cases were adult type granulosa cell tumors (GCT) and one was a juvenile type. All cases were unilateral. GCT arising from left ovary were seven, right, three. The largest diameter of the tumors ranged from 6.8 to 24 cm (mean: 11.9 cm). All had well-defined margins. Ascites was seen in four cases. Among ten cases of GCT, six were mainly solid (type 3). One case manifested as a unilocular cystic mass without mural nodule or septation. Three were multilocular cystic masses and no mural nodule was found in all three cases. Metastases to peritoneum and lymph nodes was seen in one case. The ultrasonographic findings of GCT are various but combined with clinical and laboratory findings they could be helpful in the differential diagnosis of ovarian tumors.

  19. Granulosa cell tumor of ovary: US findings

    Energy Technology Data Exchange (ETDEWEB)

    Jin, Yong Hyun; Jeon, Hae Jeong; Lee, Chang Dea; Cho, Young Kwon [Kun Kuk University School of Medicine, Seoul (Korea, Republic of); Kang, Chang Ho; Park, Yong Hyun [Korea University School of Medicine, Seoul (Korea, Republic of); Kim, Myung Gyu [Korea University School of Medicine, Seoul (Korea, Republic of); Lee, Yeon Hee [Kang Nam Cha General Hospital, Seoul (Korea, Republic of); Kim, Young Hwa; Lee, Hye Kyung [Dan Kuk University School of Medicine (Korea, Republic of)

    1999-06-15

    To describe ultrasonographic findings of ovarian granulosa cell tumor (GCT) and to determine their possible value in the differential diagnosis of ovarian tumors. Sonographic appearances of ten cases of pathologically proven GC Ts were retrospectively reviewed regarding their location, size, outer margin, the echo pattern of the tumor, endometrial thickness, presence of ascites, and metastasis to adjacent tissue or distant sites. 3.0-3.5 MHz trans-abdominal US or 5.0-6.5 MHz transvaginal US were used. The sonographic features could be classified as follows: unilocular cystic mass without nodule or septation (type 1), multilocular cystic mass (type 2), and solid mass (type 3). Pathologically nine cases were adult type granulosa cell tumors (GCT) and one was a juvenile type. All cases were unilateral. GCT arising from left ovary were seven, right, three. The largest diameter of the tumors ranged from 6.8 to 24 cm (mean: 11.9 cm). All had well-defined margins. Ascites was seen in four cases. Among ten cases of GCT, six were mainly solid (type 3). One case manifested as a unilocular cystic mass without mural nodule or septation. Three were multilocular cystic masses and no mural nodule was found in all three cases. Metastases to peritoneum and lymph nodes was seen in one case. The ultrasonographic findings of GCT are various but combined with clinical and laboratory findings they could be helpful in the differential diagnosis of ovarian tumors.

  20. Effects of progesterone and its metabolites on human granulosa cells.

    Science.gov (United States)

    Pietrowski, D; Gong, Y; Mairhofer, M; Gessele, R; Sator, M

    2014-02-01

    The corpus luteum (CL) is under control of gonadotrophic hormones and produces progesterone, which is necessary for endometrial receptivity. Recent studies have shown that progesterone and its metabolites are involved in cell proliferation and apoptosis of cancer cells. Here weanalyzed the role of progesterone and its meta-bolites on luteinized granulosa cells (LGC) by FACS analysis and quantitative Real-Time PCR. We detected the mRNA of the progesterone metabolizing genes SRD5A1, AKR1C1, and AKR1C2 in LGC. The stimulation of LGC with progesterone or progesterone metabolites did not show any effect on the mRNA expression of these genes. However, a downregulation of Fas expression was found to be accomplished by progesterone and human chorionic gonadotropin. Our findings do not support the concept of an effect of progesterone metabolites on LGCs. However, it suggests an antiapoptotic effect of hCG and progesterone during corpus luteum development by downregulation of Fas. © Georg Thieme Verlag KG Stuttgart · New York.

  1. 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

  2. The Cultivation of Human Granulosa Cells

    Directory of Open Access Journals (Sweden)

    Lenka Brůčková

    2008-01-01

    Full Text Available The major functions of granulosa cells (GCs include the production of steroids, as well as a myriad of growth factors to interact with the oocyte during its development within the ovarian follicle. Also FSH stimulates GCs to convert androgens (coming from the thecal cells to estradiol by aromatase. However, after ovulation the GCs produce progesterone that may maintain a potential pregnancy. Experiments with human GCs are mainly focused on the purification of GCs from ovarian follicular fluid followed by FACS analysis or short-term cultivation. The aim of our study was to cultivate GCs for a long period, to characterize their morphology and phenotype. Moreover, we have cultivated GCs under gonadotropin stimulation in order to simulate different pathological mechanisms during folliculogenesis (e.g. ovarian hyperstimulation syndrome. GCs were harvested from women undergoing in vitro fertilization. Complex oocyte-cumulus oophorus was dissociated by hyaluronidase. The best condition for transport of GCs was optimized as short transport in follicular fluid at 37 °C. GCs expansion medium consisted of DMEM/F12, 2 % FCS, ascorbic acid, dexamethasone, L-glutamine, gentamycine, penicillin, streptomycin and growth factors (EGF, bFGF. GCs transported in follicular fluid and cultivated in 2 % FCS containing DMEM/F12 medium supplemented with follicular fluid presented increased adhesion, proliferation, viability and decreased doubling time. Cell viability was 92 % and mean cell doubling time was 52 hrs. We have optimized transport and cultivation protocols for long-term cultivation of GCs.

  3. Prohibitin( PHB) roles in granulosa cell physiology.

    Science.gov (United States)

    Chowdhury, Indrajit; Thomas, Kelwyn; Thompson, Winston E

    2016-01-01

    Ovarian granulosa cells (GC) play an important role in the growth and development of the follicle in the process known as folliculogenesis. In the present review, we focus on recent developments in prohibitin (PHB) research in relation to GC physiological functions. PHB is a member of a highly conserved eukaryotic protein family containing the repressor of estrogen activity (REA)/stomatin/PHB/flotillin/HflK/C (SPFH) domain (also known as the PHB domain) found in diverse species from prokaryotes to eukaryotes. PHB is ubiquitously expressed in a circulating free form or is present in multiple cellular compartments including mitochondria, nucleus and plasma membrane. In mitochondria, PHB is anchored to the mitochondrial inner membrane and forms complexes with the ATPases associated with proteases having diverse cellular activities. PHB continuously shuttles between the mitochondria, cytosol and nucleus. In the nucleus, PHB interacts with various transcription factors and modulates transcriptional activity directly or through interactions with chromatin remodeling proteins. Many functions have been attributed to the mitochondrial and nuclear PHB complexes such as cellular differentiation, anti-proliferation, morphogenesis and maintenance of the functional integrity of the mitochondria. However, to date, the regulation of PHB expression patterns and GC physiological functions are not completely understood.

  4. Radiologic findings of ovarian granulosa cell tumor

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jong Chul [Chungnam National Univ. College of Medicine, Taejon (Korea, Republic of)

    1997-10-01

    To determine, through an analysis of radiologic findings, whether the findings of granulosa cell tumors (GCTs) of the ovary are specific. The radiologic findings (ultrasonography, computed tomography, and magnetic resonance imaging) of 16 pathologically proven ovarian GCTs in 15 patients were retrospectively analysed for the site of origin, staging, largest diameter, margin, solid and/or cystic components, degree of enhancement, and associated endometrial hyperplasia, ascites, and local and/or distant metastasis. Unilateral ovarian GCTs were found in 14 patients, and bilateral tumors in one. Of a total of 16 tumors, 13 were of the adult type, and three were juvenile; their largest diameter ranged from 1 to 26(mean, 15.6)cm. Eleven tumors were well-defined, two were cystic, and one small tumor was solid. Of 13 mixed tumors, three had hemorrhagic portions, and five had multilocular cystic portions. Metastases to the uterus, tubes, rectum, lymph nodes, or liver were found in six patients, and associated endometrial hyperplasia in two. Radiologically, ovarian GCTs showed well-defined or encapsulated soft tissue masses with some hemorrhagic, multilocular or focal cystic components, as well as associated endometrial thickening and local or distant metastasis. These and clinical findings may be useful in the diagnosis of ovarian GCTs.

  5. 4-Aminopyridine Decreases Progesterone Production by Porcine Granulosa Cells

    Directory of Open Access Journals (Sweden)

    Mitchell Brianna M

    2003-04-01

    Full Text Available Abstract Background Ion channels occur as large families of related genes with cell-specific expression patterns. Granulosa cells have been shown to express voltage-gated potassium channels from more than one family. The purpose of this study was to determine the effects of 4-aminopyridine (4-AP, an antagonist of KCNA but not KCNQ channels. Methods Granulosa cells were isolated from pig follicles and cultured with 4-AP, alone or in combination with FSH, 8-CPT-cAMP, estradiol 17β, and DIDS. Complimentary experiments determined the effects of 4-AP on the spontaneously established pig granulosa cell line PGC-2. Granulosa cell or PGC-2 function was assessed by radio-immunoassay of media progesterone accumulation. Cell viability was assessed by trypan blue exclusion. Drug-induced changes in cell membrane potential and intracellular potassium concentration were documented by spectrophotometric determination of DiBAC4(3 and PBFI fluorescence, respectively. Expression of proliferating cell nuclear antigen (PCNA and steroidogenic acute regulatory protein (StAR was assessed by immunoblotting. Flow cytometry was also used to examine granulosa cell viability and size. Results 4-AP (2 mM decreased progesterone accumulation in the media of serum-supplemented and serum-free granulosa cultures, but inhibited cell proliferation only under serum-free conditions. 4-AP decreased the expression of StAR, the production of cAMP and the synthesis of estradiol by PGC-2. Addition of either 8-CPT-cAMP or estradiol 17β to serum-supplemented primary cultures reduced the inhibitory effects of 4-AP. 4-AP treatment was also associated with increased cell size, increased intracellular potassium concentration, and hyperpolarization of resting membrane potential. The drug-induced hyperpolarization of resting membrane potential was prevented either by decreasing extracellular chloride or by adding DIDS to the media. DIDS also prevented 4-AP inhibition of progesterone production

  6. Control of fibronectin synthesis by rat granulosa cells in culture

    International Nuclear Information System (INIS)

    Skinner, M.K.; Dorrington, J.H.

    1984-01-01

    The secreted and cellular [ 35 S]methionine-radiolabeled proteins of cultured rat granulosa cells were separated by electrophoresis on sodium dodecylsulfate (SDS) polyacrylamide gradient slab gels. From 24 to 72 h of culture FSH increased the intensity of labeling of most of the secreted proteins. A 220,000-dalton protein, however, increased in intensity only in control cultures and became the major secreted protein after 72 h, comprising 20% of the total radiolabeled proteins. This protein was identified as fibronectin by immunoprecipitation. There was no increase in the secreted or cellular fibronectin in FSH- or testosterone- and insulin-treated cultures. These studies indicate that a component of extracellular matrix is a major secretory product of unstimulated immature granulosa cells. As hormones induce the differentiated functions of granulosa cells in culture, the secretion of fibronectin is inhibited

  7. 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.

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

    International Nuclear Information System (INIS)

    Paro, Rita; Tiboni, Gian Mario; Buccione, Roberto; Rossi, Gianna; Cellini, Valerio; Canipari, Rita; Cecconi, Sandra

    2012-01-01

    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.

  9. 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.

  10. Precocious pseudopuberty due to juvenile granulosa cell tumor

    International Nuclear Information System (INIS)

    Saeed, G.A.; Farooq, N.

    2003-01-01

    A case of precocious puberty occurring in a young girl is presented. Vaginal bleeding and secondary sexual characteristics had occurred at 7 years of age associated with an abdominal mass. These findings were due to a functional juvenile granulosa cell tumor in the right ovary. Right adenectomy was performed. Histopathology was confirmatory. (author)

  11. Hormone therapy in ovarian granulosa cell tumors: a systematic review

    NARCIS (Netherlands)

    van Meurs, Hannah S.; van Lonkhuijzen, Luc R. C. W.; Limpens, Jacqueline; van der Velden, Jacobus; Buist, Marrije R.

    2014-01-01

    This systematic review assessed the effectiveness of hormone therapy (HT) in patients with a granulosa cell tumor (GCT) of the ovary. Medline (OVID), EMBASE (OVID), the Cochrane Central Register of Controlled Trials (CENTRAL), prospective trial registers and PubMed (as supplied by publisher-subset)

  12. 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

  13. INHIBIN AS A MARKER FOR GRANULOSA-CELL TUMOR

    NARCIS (Netherlands)

    LAPPOHN, RE; BURGER, HG; BOUMA, J; BANGAH, M; KRANS, M

    1992-01-01

    In order to determine whether serum-immunoreactive inhibin could constitute a biochemical marker for the presence and progression of ovarian granulosa cell tumors and their metastases, we measured immunoreactive inhibin concentrations in series of serum samples obtained from 8 patients with

  14. Hepatic Metastases of Granulosa Cell Tumour of the Ovary

    Directory of Open Access Journals (Sweden)

    José I. Rodríguez García

    1996-01-01

    Full Text Available A case of metastatic granulosa cell tumour of the ovary is reported. Investigations revealed a secondary tumour in segment VI and VII of the liver. Right hepatic resection was performed. Microscopic findings revealed a tumour with histological features identical to that removed eleven years before.

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

    International Nuclear Information System (INIS)

    Fa, Svetlana; Pogrmic-Majkic, Kristina; Samardzija, Dragana; Glisic, Branka; Kaisarevic, Sonja; Kovacevic, Radmila; Andric, Nebojsa

    2013-01-01

    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

  16. 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

  17. 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.

  18. 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.

  19. Radiologic findings of granulosa cell tumor of the ovary

    Energy Technology Data Exchange (ETDEWEB)

    Sohn, Jung Eun; Kim, Kie Hwan; Yoo, Ji Young; Lee, Eun Chun; Lee, Tae Hyun; Chin, Soo Il [Korea Cancer Center Hospital, Seoul (Korea, Republic of)

    1997-08-01

    To evaluate the radiologic findings of granulosa cell tumor of the ovary. Fourteen cases(fifteen tumors) of pathologically confirmed ovarian granulosa cell tumor were retrospectively analyzed on the basis of CT(n=10), MR imaging(n=4), and ultrasound(n=7) findings. The patients' mean age was 44.3(range, 5-71)years. The mean diameter of the tumors was 12.1(range, 5-26.5)cm. Thirteen cases were unilateral, and one was bilateral. Eleven tumors(ten cases) were mainly solid and eight of these had focal cystic components. Multilocular cysts accounted for three cases, and in two of these, mural nodules were present. One case was a unilocular cyst with no mural nodule. Ten cases were well demarcated. All the solid tumors were enhanced on postcontrast CT and MR imaging. Endometrial thickening was seen in five cases, ascites in six, and peritoneal implants or omental fat infiltration in five. One was associated with lymph node metastasis. All the postmenopausal patients had solid tumors, whereas 66.7%(4 of 6 cases) of young adults and children had cystic tumors. Granulosa cell tumors of the ovary were solid or cystic;the former were more common. There were no characteristic findings which permitted definitive differentiation from other ovarian tumors.

  20. 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

  1. Effect of mono-(2-ethylhexyl) phthalate on steroid production of human granulosa cells

    International Nuclear Information System (INIS)

    Reinsberg, Jochen; Wegener-Toper, Petra; Ven, Katrin van der; Ven, Hans van der; Klingmueller, Dietrich

    2009-01-01

    The phthalate ester mono-(2-ethylhexyl) phthalate (MEHP) is the active metabolite of di-(2-ethylhexyl) phthalate, a high-production-volume chemical used as a plasticizer and solvent in numerous consumer products. MEHP has been demonstrated to be a reproductive toxicant in rodents decreasing estradiol and progesterone production in preovulatory granulosa cells. In the present study, we examined the effect of MEHP on steroid production of human granulosa-lutein (GL) cells. Human GL cells collected from women undergoing in vitro fertilization were cultured in medium containing FSH, hCG and 8-Br-cAMP, respectively, together with various concentrations of MEHP (0-500 μmol L -1 ). After incubation for 48 h estradiol and progesterone were assayed in the spent culture medium. Furthermore, aromatase activity and mRNA levels of GL cells were determined. Basal as well as FSH-, hCG- and 8-Br-cAMP-stimulated estradiol production of GL cells was suppressed by MEHP in a dose-dependent manner (IC 50 = 105 μmol L -1 , 138 μmol L -1 , 49 μmol L -1 and 78 μmol L -1 ). Furthermore aromatase activity and mRNA levels were reduced in GL cells cultured with MEHP. In contrast, MEHP did not alter the production of progesterone up to a concentration of 167 μmol L -1 . The present data indicate that MEHP is a specific inhibitor of estradiol production in human GL cells with a post-cAMP site of action. The inhibition of estradiol production obviously results from a reduction of aromatase activity on the transcript level. As the in vitro effective doses of MEHP are within the range of real environmental exposure levels an inhibitory effect on estrogen production in vivo seems to be possible.

  2. Activin A, B and AB decrease progesterone production by down-regulating StAR in human granulosa cells.

    Science.gov (United States)

    Chang, Hsun-Ming; Cheng, Jung-Chien; Huang, He-Feng; Shi, Feng-Tao; Leung, Peter C K

    2015-09-05

    Activins are homo- or heterodimers of inhibin β subunits that play important roles in the reproductive system. Our previous work has shown that activins A (βAβA), B (βBβB) and AB (βAβB) induce aromatase/estradiol, but suppress StAR/progesterone production in human granulosa-lutein cells. However, the underlying molecular determinants of these effects have not been examined. In this continuing study, we used immortalized human granulosa cells (SVOG) to investigate the effects of activins in regulating StAR/progesterone and the potential mechanisms of action. In SVOG cells, activins A, B and AB produced comparable down-regulation of StAR expression and progesterone production. In addition, all three activin isoforms induced equivalent phosphorylation of both SMAD2 and SMAD3. Importantly, the activin-induced down-regulation of StAR, increase in SMAD2/3 phosphorylation, and decrease in progesterone were abolished by the TGF-β type I receptor inhibitor SB431542. Interestingly, the small interfering RNA-mediated knockdown of ALK4 but not ALK5 reversed the activin-induced suppression of StAR. Furthermore, the knockdown of SMAD4 or SMAD2 but not SMAD3 abolished the inhibitory effects of all three activin isoforms on StAR expression. These results provide evidence that activins A, B and AB down-regulate StAR expression and decrease progesterone production in human granulosa cells, likely via an ALK4-mediated SMAD2/SMAD4-dependent pathway. Our findings provide important insights into the molecular mechanisms underlying the regulatory effects of activins on human granulosa cell steroidogenesis. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

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

    Directory of Open Access Journals (Sweden)

    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

  4. Evidence for alternative pathways of granulosa cell death in healthy and slightly atretic bovine antral follicles.

    Science.gov (United States)

    Van Wezel, I L; Dharmarajan, A M; Lavranos, T C; Rodgers, R J

    1999-06-01

    Granulosa cell death is an early feature of atresia; however, there are many apparent contradictions in the literature concerning the mode of granulosa cell death. We have therefore examined this process in bovine healthy and atretic antral follicles, using a variety of established techniques. Light and electron microscopic observations indicated the presence of pyknotic or shrunken nuclei in both the membrana granulosa and the antrum. In the membrana granulosa, these nuclei were frequently crescent shaped and uniformly electron dense and were approximately the same size as healthy nuclei, all of which are typical of early apoptosis. However, these nuclei were within the membranes of a healthy granulosa cell, suggesting that phagocytosis by a neighboring granulosa cell is an unusually early event in the apoptotic pathway of granulosa cells. In the membrana granulosa, pyknotic nuclei stained intensely with hematoxylin but weakly with the DNA-intercalating stain propidium iodide. A percentage of these pyknotic nuclei stained by TUNEL (terminal deoxy-UTP nick end-labeling). However, in the antrum, the pyknotic nuclei and larger globules of DNA stained intensely with both hematoxylin and propidium iodide, but were not TUNEL positive. The comet assay of cell death produced a streak tail of randomly nicked DNA, rather than the plume of low mol wt apoptotic DNA. Globules collected from fresh follicular fluid stained intensely with propidium iodide and were shown by PAGE to contain DNA, the majority of which was high mol wt. In conclusion, granulosa cells within the membrana granulosa die by apoptosis, with phagocytosis by a neighboring cell preceding any potential budding of the nucleus or cell itself. Granulosa cells near the antrum are sloughed off into the antrum, and their death has features more consistent with that of other cell types that undergo death as a result of terminal differentiation.

  5. Direct antigonadal activity of cannabinoids: suppression of rat granulosa cell functions.

    Science.gov (United States)

    Adashi, E Y; Jones, P B; Hsueh, A J

    1983-02-01

    The direct effects of delta 9-tetrahydrocannabinol (THC) and related cannabinoids on ovarian granulosa cells were studied in vitro. Granulosa cells from immature, hypophysectomized, estrogen-treated rats were cultured for 2 days in an androstenedione-supplemented medium in the presence or absence of follicle-stimulating hormone (FSH) (10 ng/ml) with or without cannabinoids. FSH treatment increased progesterone and estrogen biosynthesis, whereas concomitant treatment with THC led to a dose-dependent inhibition of the FSH-stimulated accumulation of progesterone and estrogen with ED50 values of 3.5 +/- 0.3 X 10(-7) and 1.8 +/- 0.2 X 10(-6) M, respectively. Treatment with related but nonpsychoactive cannabinoids (cannabidiol, cannabinol, cannabigerol, or cannabichromene) was equally effective. The THC-induced inhibition of progesterone production was reversible and was associated with an inhibition of pregnenolone biosynthesis and a decrease of 3 beta-hydroxysteroid dehydrogenase activity. In addition, treatment with THC brought about a dose-dependent inhibition of the FSH-induced increase in luteinizing hormone (LH) receptors. The inhibitory effects of THC were not associated with changes in cell number, protein content, or cell viability. Thus, THC exerts direct inhibitory effects on FSH-dependent functions related to steroidogenesis and the acquisition of LH receptors, all of which are essential to follicular maturation. Because plasma concentrations of THC similar to those used in this study have been reported in human beings, repeated exposure of female users to THC may lead to ovarian dysfunction, due in part, to the direct antigonadal activity to THC.

  6. First case of juvenile granulosa cell tumor in an adult with Ollier disease.

    NARCIS (Netherlands)

    Rietveld, L.; Nieboer, T.E.; Kluivers, K.B.; Schreuder, H.W.B.; Bulten, J.; Massuger, L.F.A.G.

    2009-01-01

    Ollier's disease (OD) is a rare disorder associated with the presence of multiple enchondromas. Granulosa cell tumors are rare sex cord-stromal ovarian tumors. This is the first report of a patient in her fourth decade with a combination of OD and juvenile granulosa cell tumor.A 36-year-old woman

  7. OAZ1 knockdown enhances viability and inhibits ER and LHR transcriptions of granulosa cells in geese.

    Directory of Open Access Journals (Sweden)

    Bo Kang

    Full Text Available An increasing number of studies suggest that ornithine decarboxylase antizyme 1 (OAZ1, which is regarded as a tumor suppressor gene, regulates follicular development, ovulation, and steroidogenesis. The granulosa cells in the ovary play a critical role in these ovarian functions. However, the action of OAZ1 mediating physiological functions of granulosa cells is obscure. OAZ1 knockdown in granulosa cells of geese was carried out in the current study. The effect of OAZ1 knockdown on polyamine metabolism, cell proliferation, apoptosis, and hormone receptor transcription of primary granulosa cells in geese was measured. The viability of granulosa cells transfected with the shRNA OAZ1 at 48 h was significantly higher than the control (p<0.05. The level of putrescine and spermidine in granulosa cells down-regulating OAZ1 was 7.04- and 2.11- fold higher compared with the control, respectively (p<0.05. The CCND1, SMAD1, and BCL-2 mRNA expression levels in granulosa cells down-regulating OAZ1 were each significantly higher than the control, respectively (p<0.05, whereas the PCNA and CASPASE 3 expression levels were significantly lower than the control (p<0.05. The estradiol concentration, ER and LHR mRNA expression levels were significantly lower in granulosa cells down-regulating OAZ1 compared with the control (p<0.05. Taken together, our results indicated that OAZ1 knockdown elevated the putrescine and spermidine contents and enhanced granulosa cell viability and inhibited ER and LHR transcriptions of granulosa cells in geese.

  8. BMP15 suppresses progesterone production by down-regulating StAR via ALK3 in human granulosa cells.

    Science.gov (United States)

    Chang, Hsun-Ming; Cheng, Jung-Chien; Klausen, Christian; Leung, Peter C K

    2013-12-01

    In addition to somatic cell-derived growth factors, oocyte-derived growth differentiation factor (GDF)9 and bone morphogenetic protein (BMP)15 play essential roles in female fertility. However, few studies have investigated their effects on human ovarian steroidogenesis, and fewer still have examined their differential effects or underlying molecular determinants. In the present study, we used immortalized human granulosa cells (SVOG) and human granulosa cell tumor cells (KGN) to compare the effects of GDF9 and BMP15 on steroidogenic enzyme expression and investigate potential mechanisms of action. In SVOG cells, neither GDF9 nor BMP15 affects the mRNA levels of P450 side-chain cleavage enzyme or 3β-hydroxysteroid dehydrogenase. However, treatment with BMP15, but not GDF9, significantly decreases steroidogenic acute regulatory protein (StAR) mRNA and protein levels as well as progesterone production. These suppressive effects, along with the induction of Sma and Mad-related protein (SMAD)1/5/8 phosphorylation, are attenuated by cotreatment with 2 different BMP type I receptor inhibitors (dorsomorphin and DMH-1). Furthermore, depletion of activin receptor-like kinase (ALK)3 using small interfering RNA reverses the effects of BMP15 on SMAD1/5/8 phosphorylation and StAR expression. Similarly, knockdown of ALK3 abolishes BMP15-induced SMAD1/5/8 phosphorylation in KGN cells. These results provide evidence that oocyte-derived BMP15 down-regulates StAR expression and decreases progesterone production in human granulosa cells, likely via ALK3-mediated SMAD1/5/8 signaling. Our findings suggest that oocyte may play a critical role in the regulation of progesterone to prevent premature luteinization during the late stage of follicle development.

  9. Regulation of MMP2 and MMP9 metalloproteinases by FSH and growth factors in bovine granulosa cells

    Directory of Open Access Journals (Sweden)

    Valerio M. Portela

    2009-01-01

    Full Text Available Matrix metalloproteinases (MMP are key enzymes involved in tissue remodeling. Within the ovary, they are believed to play a major role in ovulation, and have been linked to follicle atresia. To gain insight into the regulation of MMPs, we measured the effect of hormones and growth factors on MMP2 and MMP9 mRNA levels in non-luteinizing granulosa cells in serum-free culture. FSH and IGF1 both stimulated estradiol secretion and inhibited MMP2 and MMP9 mRNA abundance. In contrast, EGF and FGF2 both inhibited estradiol secretion but had no effect on MMP expression. At physiological doses, none of these hormones altered the proportion of dead cells. Although we cannot link MMP expression with apoptosis, the specific down regulation by the gonadotropic hormones FSH and IGF1 in vitro suggests that excess MMP2 and MMP9 expression is neither required nor desired for follicle development.

  10. Lactate promotes specific differentiation in bovine granulosa cells depending on lactate uptake thus mimicking an early post-LH stage

    Directory of Open Access Journals (Sweden)

    Anja Baufeld

    2018-02-01

    Full Text Available Abstract Background The LH-induced folliculo-luteal transformation is connected with alterations of the gene expression profile in cells of the granulosa layer. It has been described that hypoxic conditions occur during luteinization, thus favoring the formation of L-lactate within the follicle. Despite being a product of anaerobic respiration, L-lactate has been shown to act as a signaling molecule affecting gene expression in neuronal cells. During the present study, we tested the hypothesis that L-lactate may influence differentiation of follicular granulosa cells (GC. Methods In a bovine granulosa cell culture model effects of L- and D-lactate, of increased glucose concentrations and of the lactate transport inhibitor UK5099 were analyzed. Steroid hormone production was analyzed by RIA and the abundance of key transcripts was determined by quantitative real-time RT-PCR. Results L-lactate decreased the production of estradiol and significantly affected selected genes of the folliculo-luteal transition as well as genes of the lactate metabolism. CYP19A1, FSHR, LHCGR were down-regulated, whereas RGS2, VNN2, PTX3, LDHA and lactate transporters were up-regulated. These effects could be partly or completely reversed by pre-treatment of the cells with UK5099. The non-metabolized enantiomer D-lactate had even more pronounced effects on gene expression, whereas increased glucose concentrations did not affect transcript abundance. Conclusions In summary, our data suggest that L-lactate specifically alters physiological and molecular characteristics of GC. These effects critically depend on L-lactate uptake, but are not triggered by increased energy supply. Further, we could show that L-lactate has a positive feedback on the lactate metabolism. Therefore, we hypothesize that L-lactate acts as a signaling molecule in bovine and possibly other monovular species supporting differentiation during the folliculo-luteal transformation.

  11. Pathogenesis and treatment of adult-type granulosa cell tumor of the ovary.

    Science.gov (United States)

    Färkkilä, Anniina; Haltia, Ulla-Maija; Tapper, Johanna; McConechy, Melissa K; Huntsman, David G; Heikinheimo, Markku

    2017-08-01

    Adult-type granulosa cell tumor is a clinically and molecularly unique subtype of ovarian cancer. These tumors originate from the sex cord stromal cells of the ovary and represent 3-5% of all ovarian cancers. The majority of adult-type granulosa cell tumors are diagnosed at an early stage with an indolent prognosis. Surgery is the cornerstone for the treatment of both primary and relapsed tumor, while chemotherapy is applied only for advanced or non-resectable cases. Tumor stage is the only factor consistently associated with prognosis. However, every third of the patients relapse, typically in 4-7 years from diagnosis, leading to death in 50% of these patients. Anti-Müllerian Hormone and inhibin B are currently the most accurate circulating biomarkers. Adult-type granulosa cell tumors are molecularly characterized by a pathognomonic somatic missense point mutation 402C->G (C134W) in the transcription factor FOXL2. The FOXL2 402C->G mutation leads to increased proliferation and survival of granulosa cells, and promotes hormonal changes. Histological diagnosis of adult-type granulosa cell tumor is challenging, therefore testing for the FOXL2 mutation is crucial for differential diagnosis. Large international collaborations utilizing molecularly defined cohorts are essential to improve and validate new treatment strategies for patients with high-risk or relapsed adult-type granulosa cell tumor. Key Messages: Adult-type granulosa cell tumor is a unique ovarian cancer with an indolent, albeit unpredictable disease course. Adult-type granulosa cell tumors harbor a pathognomonic somatic missense mutation in transcription factor FOXL2. The key challenges in the treatment of patients with adult-type granulosa cell tumor lie in the identification and management of patients with high-risk or relapsed disease.

  12. Removal of glycosaminoglycans from bovine granulosa cells contributes to increased binding of hydrogen-3 heparin

    Energy Technology Data Exchange (ETDEWEB)

    Ax, R.L.; Stodd, C.M.; Boehm, S.K.; Bellin, M.E.

    1986-02-01

    Granulosa cells from small or large bovine follicles were pretreated with enzymes that hydrolyze various glycosaminoglycans, and binding of (/sup 3/H)-heparin to the granulosa was measured. Binding of (/sup 3/H) heparin increased significantly after enzymatic pretreatments with chondroitinase ABC and fungal hyaluronidase, and similar results were obtained with granulosa from small and large follicles. No changes in binding of (/sup 3/H) heparin were detected after hydrolyses with chondroitinase AC and heparinase in either follicle size. Heparitinase, which hydrolyzes heparan sulfate, led to a significant 50% increase in binding of (/sup 3/H) heparin to granulosa from large follicles but was without effect in small follicles. These results suggest that the lower binding of (/sup 3/H) heparin, which has been reported with follicular enlargement, may be due to heparan sulfate occupying or obstructing binding sites for heparin on granulosa from large follicles.

  13. Testicular juvenile granulosa cell tumor in a newborn: case report and review of the literature.

    Science.gov (United States)

    Alexiev, Borislav A; Alaish, Samuel M; Sun, Chen-Chih

    2007-07-01

    Juvenile granulosa cell tumor of the testis of neonates and infants is an uncommon lesion frequently associated with abnormal sex chromosome and ambiguous genitalia. This report describes a juvenile granulosa cell tumor arising in the testis of a neonate. Chromosome analysis of the tumor showed a normal male karyotype 46 XY. Histopathology and immunohistochemical studies revealed the occurrence of 2 well-differentiated epithelial-like and smooth muscle-like components in the neoplasm. The morphologic clues leading to the correct diagnosis of juvenile granulosa cell tumor and the possible histogenesis are briefly discussed.

  14. Cellular and exosome mediated molecular defense mechanism in bovine granulosa cells exposed to oxidative stress.

    Directory of Open Access Journals (Sweden)

    Mohammed Saeed-Zidane

    Full Text Available Various environmental insults including diseases, heat and oxidative stress could lead to abnormal growth, functions and apoptosis in granulosa cells during ovarian follicle growth and oocyte maturation. Despite the fact that cells exposed to oxidative stress are responding transcriptionally, the potential release of transcripts associated with oxidative stress response into extracellular space through exosomes is not yet determined. Therefore, here we aimed to investigate the effect of oxidative stress in bovine granulosa cells in vitro on the cellular and exosome mediated defense mechanisms. Bovine granulosa cells were aspirated from ovarian follicles and cultured in DMEM/F-12 Ham culture medium supplemented with 10% exosome-depleted fetal bovine serum. In the first experiment sub-confluent cells were treated with 5 μM H2O2 for 40 min to induce oxidative stress. Thereafter, cells were subjected to ROS and mitochondrial staining, cell proliferation and cell cycle assays. Furthermore, gene and protein expression analysis were performed in H2O2-challenged versus control group 24 hr post-treatment using qRT-PCR and immune blotting or immunocytochemistry assay, respectively. Moreover, exosomes were isolated from spent media using ultracentrifugation procedure, and subsequently used for RNA isolation and qRT-PCR. In the second experiment, exosomes released by granulosa cells under oxidative stress (StressExo or those released by granulosa cells without oxidative stress (NormalExo were co-incubated with bovine granulosa cells in vitro to proof the potential horizontal transfer of defense molecules from exosomes to granulosa cells and investigate any phenotype changes. Exposure of bovine granulosa cells to H2O2 induced the accumulation of ROS, reduced mitochondrial activity, increased expression of Nrf2 and its downstream antioxidant genes (both mRNA and protein, altered the cell cycle transitions and induced cellular apoptosis. Granulosa cells

  15. Fetuin and fetuin messenger RNA in granulosa cells of the rat ovary

    DEFF Research Database (Denmark)

    Høyer, Poul Erik; Terkelsen, O B; Grete Byskov, A

    2001-01-01

    during maturation of the oocyte. We demonstrated fetuin mRNA in the rat ovary by reverse transcriptase-polymerase chain reaction and localized it by in situ hybridization. Fetuin mRNA was present in all granulosa cells of growing and large follicles. Immunohistochemical analysis revealed that the fetuin...... protein was only present in some of the small, growing follicles. In large, healthy follicles, fetuin protein was confined to cumulus cells and granulosa cells bordering the antrum. Fetuin was present in atretic follicles, but the staining pattern differed from that of healthy follicles. The follicular...... antrum contained a substantial amount of fetuin, but whether granulosa cells secreted it or it originated in the ovarian blood supply could not be confirmed. We concluded that at least a portion of the fetuin is produced by granulosa cells of growing and large follicles, suggesting that fetuin may...

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

    International Nuclear Information System (INIS)

    Gao, Hui; Lin, Lu; Haq, Ihtesham Ul; Zeng, Shen-ming

    2016-01-01

    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.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

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

    Directory of Open Access Journals (Sweden)

    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.

  19. Effects of an inhibitor of the γ-secretase complex on proliferation and apoptotic parameters in a FOXL2-mutated granulosa tumor cell line (KGN).

    Science.gov (United States)

    Irusta, Griselda; Pazos, Maria Camila; Maidana, Camila Pazos; Abramovich, Dalhia; De Zúñiga, Ignacio; Parborell, Fernanda; Tesone, Marta

    2013-07-01

    Ovarian granulosa cell tumors (GCTs) represent 3%-5% of all ovarian malignancies. Treatments have limited proven efficacy and biologically targeted treatment is lacking. The aim of this study was to investigate the role of Notch signaling in the proliferation, steroidogenesis, apoptosis, and phosphatidylinositol 3-kinase (PI3K)/AKT pathway in a FOXL2-mutated granulosa tumor cell line (KGN) representative of the adult form of GCTs. When Notch signaling is initiated, the receptors expose a cleavage site in the extracellular domain to the metalloproteinase TACE and, following this cleavage, Notch undergoes another cleavage mediated by the presenilin-gamma-secretase complex. To achieve our goal, DAPT, an inhibitor of the gamma-secretase complex, was used to investigate the role of the Notch system in parameters associated with cell growth and death, using a human granulosa cell tumor line (KGN) as an experimental model. We observed that JAGGED1, DLL4, NOTCH1, and NOTCH4 were highly expressed in KGN cells as compared to granulosa-lutein cells obtained from assisted reproductive techniques patients. The proliferation and viability of KGN cells, as well as progesterone and estradiol production, decreased in the presence of 20 μM DAPT. Apoptotic parameters like PARP and caspase 8 cleavages, BAX, and BCLXs increased in KGN cells cultured with DAPT, whereas others such as BCL2, BCLXl, FAS, and FAS ligand did not change. AKT phosphorylation decreased and PTEN protein increased when Notch signaling was inhibited in KGN cells. We conclude that the Notch system acts as a survival pathway in KGN cells, and might be interacting with the PI3K/AKT pathway.

  20. PMS2 gene mutation results in DNA mismatch repair system failure in a case of adult granulosa cell tumor

    OpenAIRE

    Wang, Wen-Chung; Lee, Ya-Ting; Lai, Yen-Chein

    2017-01-01

    Background Granulosa cell tumors are rare ovarian malignancies. Their characteristics include unpredictable indolent growth with malignant potential and late recurrence. Approximately 95% are of adult type. Recent molecular studies have characterized the FOXL2 402C?>?G mutation in adult granulosa cell tumor. Our previous case report showed that unique FOXL2 402C?>?G mutation and defective DNA mismatch repair system are associated with the development of adult granulosa cell tumor. Findings In...

  1. Dual effect of insulin resistance and cadmium on human granulosa cells - In vitro study

    Energy Technology Data Exchange (ETDEWEB)

    Belani, Muskaan, E-mail: muskaanbelani@gmail.com [Department of Biochemistry, Faculty of Science, The Maharaja Sayajirao University of Baroda, Vadodara 390 002, Gujarat, India. (India); Shah, Preeti, E-mail: preeti.shah@novaivifertility.com [Nova IVI Fertility, Behind Xavier' s Ladies Hostel, 108, Swastik Society Rd., Navrangpura, Ahmedabad 390009, Gujarat, India. (India); Banker, Manish, E-mail: manish.banker@novaivifertility.com [Nova IVI Fertility, Behind Xavier' s Ladies Hostel, 108, Swastik Society Rd., Navrangpura, Ahmedabad 390009, Gujarat, India. (India); Gupta, Sarita, E-mail: saritagupta9@gmail.com [Department of Biochemistry, Faculty of Science, The Maharaja Sayajirao University of Baroda, Vadodara 390 002, Gujarat, India. (India)

    2016-12-15

    Combined exposure of cadmium (Cd) and insulin resistance (IR) might be responsible for subfertility. In the present study, we investigated the effects of Cd in vitro in IR human granulosa cells. Isolated human granulosa cells from control and polycystic ovary syndrome (PCOS) follicular fluid samples were confirmed for IR by decrease in protein expression of insulin receptor-β. Control and IR human granulosa cells were then incubated with or without 32 μM Cd. The combined effect of IR with 32 μM Cd in granulosa cells demonstrated significant decrease in expression of StAR, CYP11A1, CYP19A1, 17β-HSD, 3β-HSD, FSH-R and LH-R. Decrease was also observed in progesterone and estradiol concentrations as compared to control. Additionally, increase in protein expression of cleaved PARP-F2, active caspase-3 and a positive staining for Annexin V and PI indicated apoptosis as the mode of increased cell death ultimately leading to decreased steroidogenesis, as observed through the combined exposure. Taken together the results suggest decrease in steroidogenesis ultimately leading to abnormal development of the follicle thus compromising fertility at the level of preconception. - Highlights: • Protein expression of INSR-β in granulosa cells to differentiate PCOS-IR and NIR • Cd and IR together decrease steroidogenesis in human granulosa cells in vitro. • Cd and IR increase human granulosa cell death by increase in apoptosis. • Environment and life style are set to hamper pregnancies at preconception level.

  2. Dual effect of insulin resistance and cadmium on human granulosa cells - In vitro study

    International Nuclear Information System (INIS)

    Belani, Muskaan; Shah, Preeti; Banker, Manish; Gupta, Sarita

    2016-01-01

    Combined exposure of cadmium (Cd) and insulin resistance (IR) might be responsible for subfertility. In the present study, we investigated the effects of Cd in vitro in IR human granulosa cells. Isolated human granulosa cells from control and polycystic ovary syndrome (PCOS) follicular fluid samples were confirmed for IR by decrease in protein expression of insulin receptor-β. Control and IR human granulosa cells were then incubated with or without 32 μM Cd. The combined effect of IR with 32 μM Cd in granulosa cells demonstrated significant decrease in expression of StAR, CYP11A1, CYP19A1, 17β-HSD, 3β-HSD, FSH-R and LH-R. Decrease was also observed in progesterone and estradiol concentrations as compared to control. Additionally, increase in protein expression of cleaved PARP-F2, active caspase-3 and a positive staining for Annexin V and PI indicated apoptosis as the mode of increased cell death ultimately leading to decreased steroidogenesis, as observed through the combined exposure. Taken together the results suggest decrease in steroidogenesis ultimately leading to abnormal development of the follicle thus compromising fertility at the level of preconception. - Highlights: • Protein expression of INSR-β in granulosa cells to differentiate PCOS-IR and NIR • Cd and IR together decrease steroidogenesis in human granulosa cells in vitro. • Cd and IR increase human granulosa cell death by increase in apoptosis. • Environment and life style are set to hamper pregnancies at preconception level.

  3. Changes in keratin 8/18 expression in human granulosa cell lineage are associated to cell death/survival events: potential implications for the maintenance of the ovarian reserve.

    Science.gov (United States)

    Gaytan, F; Morales, C; Roa, J; Tena-Sempere, M

    2018-04-01

    Is keratin 8/18 (K8/K18) expression linked to cell death/survival events in the human granulosa cell lineage? A close association exists between changes in K8/K18 expression and cell death/survival events along the human granulosa cell lineage lifespan. In addition to their structural and mechanical functions, K8/K18 play essential roles regulating cell death, survival and differentiation in several non-gonadal epithelial tissues. Transfection of the granulosa-like tumor KGN cells with siRNA to interfere KRT8 and KRT18 expression increases FAS-mediated apoptosis, while an inverse association between K8/K18 expression and cell death has been found in the bovine antral follicles and corpus luteum. Yet, only fragmentary and inconclusive information exists regarding K8/K18 expression in the human ovary. Expression of K8/K18 was assessed by immunohistochemistry at different stages of the granulosa cell lineage, from flattened granulosa cells in primordial follicles to fully luteinized granulosa-lutein cells in the corpus luteum (including corpus luteum of pregnancy). Immunohistochemical detection of K8/K18 was conducted in 40 archival ovarian samples from women aged 17-39 years. K8/K18 expression was analyzed at the different stages of follicle development and corpus luteum lifespan. The proportions of primordial follicles showing all K8/K18-positive, all K8/K18 negative, or a mixture of K8/K18 negative and positive granulosa cells were quantified in 18 ovaries, divided into three age groups: ≤ 25 years (N = 6), 26-30 (N = 6) and 31-36 (N = 6) years. A total number of 1793 primordial, 750 transitional and 140 primary follicles were scored. A close association was found between changes in K8/K18 expression and cell death/cell survival events in the human granulosa cell lineage. Large secondary and early antral follicles (most of them undergoing atresia) and regressing corpora lutea displayed low/absent K8/K18 expression. Conversely, early growing and some large antral

  4. Insulin resistance enhances the mitogen-activated protein kinase signaling pathway in ovarian granulosa cells.

    Directory of Open Access Journals (Sweden)

    Linghui Kong

    Full Text Available The ovary is the main regulator of female fertility. Granulosa cell dysfunction may be involved in various reproductive endocrine disorders. Here we investigated the effect of insulin resistance on the metabolism and function of ovarian granulosa cells, and dissected the functional status of the mitogen-activated protein kinase signaling pathway in these cells. Our data showed that dexamethasone-induced insulin resistance in mouse granulosa cells reduced insulin sensitivity, accompanied with an increase in phosphorylation of p44/42 mitogen-activated protein kinase. Furthermore, up-regulation of cytochrome P450 subfamily 17 and testosterone and down-regulation of progesterone were observed in insulin-resistant mouse granulosa cells. Inhibition of p44/42 mitogen-activated protein kinase after induction of insulin resistance in mouse granulosa cells decreased phosphorylation of p44/42 mitogen-activated protein kinase, downregulated cytochrome P450 subfamily 17 and lowered progesterone production. This insulin resistance cell model can successfully demonstrate certain mechanisms such as hyperandrogenism, which may inspire a new strategy for treating reproductive endocrine disorders by regulating cell signaling pathways.

  5. Morphology and steroidogenesis of cultured granulosa cells obtained from ovaries of women treated with ionizing radiation

    International Nuclear Information System (INIS)

    Skrzypczak, J.

    1997-01-01

    The object of the study was the morphology and steroidogenesis of cultured granulosa cells obtained from 6 women aged 28-39 years who, because of Ib cervix carcinoma, were treated with ionizing radiation and later underwent surgery. It was observed that the granulosa cells were viable, had strong proliferative ability, and formed a monolayer on day 2 of culture. Contrary to our expectations, these cells produced larger amounts of steroids in culture than the control cells harvested from normal ovaries in late follicular phase. It was also found that the cells treated with ionizing radiation responded to exogenous gonadotropins with higher production of progesterone and estradiol than the controls. It is concluded that the increase in metabolic activity by granulosa cells from ovaries which had been indirectly affected by ionizing radiation is manifested by the stimulating influence of radiation on steroidogenesis. (author)

  6. Local effect of bisphenol A on the estradiol synthesis of ovarian granulosa cells from PCOS.

    Science.gov (United States)

    Wang, Yuan; Zhu, Qinling; Dang, Xuan; He, Yaqiong; Li, Xiaoxue; Sun, Yun

    2017-01-01

    Close relationship between polycystic ovary syndrome (PCOS) and bisphenol A (BPA) has drawn much attention in recent years, while the underlying mechanisms are poorly understood. In our study, we aim to detect BPA concentration in the follicular fluid and investigate its effect on estradiol synthesis in human granulosa cells from PCOS and non-PCOS patients. Follicular fluid and granulosa cells were collected from women who underwent controlled ovarian stimulation for in vitro fertilization or intracytoplasmic sperm injection. BPA concentration in the follicular fluid from PCOS patients (440.50 ± 63.70 pg/ml) was significantly higher than that from non-PCOS patients (338.00 ± 57.88 pg/ml). Expression of aromatase and estradiol synthesis in cultured granulosa cells was examined after treatment with BPA from 0.01 to 1 μM for 24 h. Expression of aromatase and estradiol synthesis was downregulated by BPA in a dose-dependent manner in PCOS, but no effect was observed in granulosa cells from non-PCOS patients. These findings provide evidence that increased BPA concentration in the follicular fluid of PCOS patients may play an important role in its pathogenesis by attenuating the expression of aromatase in granulosa cells.

  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. Paracrine effects of oocyte secreted factors and stem cell factor on porcine granulosa and theca cells in vitro

    Directory of Open Access Journals (Sweden)

    Webb Bob

    2003-08-01

    Full Text Available Abstract Oocyte control of granulosa and theca cell function may be mediated by several growth factors via a local feedback loop(s between these cell types. This study examined both the role of oocyte-secreted factors on granulosa and thecal cells, cultured independently and in co-culture, and the effect of stem cell factor (SCF; a granulosa cell derived peptide that appears to have multiple roles in follicle development. Granulosa and theca cells were isolated from 2–6 mm healthy follicles of mature porcine ovaries and cultured under serum-free conditions, supplemented with: 100 ng/ml LR3 IGF-1, 10 ng/ml insulin, 100 ng/ml testosterone, 0–10 ng/ml SCF, 1 ng/ml FSH (granulosa, 0.01 ng/ml LH (theca or 1 ng/ml FSH and 0.01 ng/ml LH (co-culture and with/without oocyte conditioned medium (OCM or 5 oocytes. Cells were cultured in 96 well plates for 144 h, after which viable cell numbers were determined. Medium was replaced every 48 h and spent medium analysed for steroids. Oocyte secreted factors were shown to stimulate both granulosa cell proliferation (P

  9. Effects of luteinizing hormone and human chorionic gonadotropin on corpus luteum cells in a spheroid cell culture system.

    Science.gov (United States)

    Walz, A; Keck, C; Weber, H; Kissel, C; Pietrowski, D

    2005-09-01

    The human corpus luteum (CL) is a highly vascularized, temporarily active endocrine gland and consists mainly of granulosa cells (GCs), theca cells (TCs), and endothelial cells (ECs). Its cyclic growth and development takes place under the influence of gonadotropic hormones. If pregnancy does occur, human chorionic gonadotropin (hCG) takes over the function of luteinizing hormone (LH) and, in contrast to LH, extends the functional life span of the CL. In this study, we investigated the effects of hCG and LH in a spheroidal cell culture model of CL development. Our data indicate that GCs secrete factors under the control of hCG that increase sprout formation of EC-spheroids. We demonstrate that the most prominent of these factors is VEGF-A. Furthermore, we found that both LH and hCG decrease sprout formation of GC-spheroids. After forming EC-GC coculture spheroids and consequently bringing GCs and ECs in close contact, sprouting increased under the influence of hCG, however not under LH. These experiments provide evidence for an hCG dependent functional switch in the GCs after coming in contact with ECs. Moreover, it demonstrates the considerably different effects of hCG and LH on GCs although their signaling is transmitted via the same receptor.

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

    International Nuclear Information System (INIS)

    Miyake, Seiji; Kobayashi, Saori; Tsubota, Kazuo; Ozawa, Yoko

    2014-01-01

    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

  11. 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.

  12. Transcriptome comparisons identify new cell markers for theca interna and granulosa cells from small and large antral ovarian follicles.

    Directory of Open Access Journals (Sweden)

    Nicholas Hatzirodos

    Full Text Available In studies using isolated ovarian granulosa and thecal cells it is important to assess the degree of cross contamination. Marker genes commonly used for granulosa cells include FSHR, CYP19A1 and AMH while CYP17A1 and INSL3 are used for thecal cells. To increase the number of marker genes available we compared expression microarray data from isolated theca interna with that from granulosa cells of bovine small (n = 10 for both theca and granulosa cells; 3-5 mm and large (n = 4 for both theca and granulosa cells, > 9 mm antral follicles. Validation was conducted by qRT-PCR analyses. Known markers such as CYP19A1, FSHR and NR5A2 and another 11 genes (LOC404103, MGARP, GLDC, CHST8, CSN2, GPX3, SLC35G1, CA8, CLGN, FAM78A, SLC16A3 were common to the lists of the 50 most up regulated genes in granulosa cells from both follicle sizes. The expression in theca interna was more consistent than in granulosa cells between the two follicle sizes. Many genes up regulated in theca interna were common to both sizes of follicles (MGP, DCN, ASPN, ALDH1A1, COL1A2, FN1, COL3A1, OGN, APOD, COL5A2, IGF2, NID1, LHFP, ACTA2, DUSP12, ACTG2, SPARCL1, FILIP1L, EGFLAM, ADAMDEC1, HPGD, COL12A1, FBLN5, RAMP2, COL15A1, PLK2, COL6A3, LOXL1, RARRES1, FLI1, LAMA2. Many of these were stromal extracellular matrix genes. MGARP, GLDC, CHST8, GPX3 were identified as new potential markers for granulosa cells, while FBLN5, OGN, RAMP2 were significantly elevated in the theca interna.

  13. Fractalkine is expressed in the human ovary and increases progesterone biosynthesis in human luteinised granulosa cells

    Directory of Open Access Journals (Sweden)

    Yan Jie

    2011-06-01

    Full Text Available Abstract Background Recent evidence from rodent ovaries has demonstrated expression of fractalkine and the existence of fractalkine receptor, and showed that there is a significant increase in steroidogenesis in response to fractalkine, yet the role of fractalkine and CX3CR1 in the human ovary is still unknown. This study aimed to determine the expression levels of fractalkine and CX3CR1 in the human ovary and to investigate their roles in sexual hormone biosynthesis by human luteinising granulosa cells. This is the first detailed report of fractalkine and CX3CR1 expression and function in the human ovary. Methods Fractalkine and CX3CR1 expression levels were measured by immunohistochemistry using ovarian tissue from pathological specimens from five individuals. Granulosa cells were obtained from patients during IVF treatment. They were cultured and treated with increasing doses of hCG with or without fractalkine. Media were collected to detect estradiol and progesterone by chemiluminescence. StAR, 3-βHSD and CYP11A expression were determined in granulosa cells treated with or without fractalkine by real-time RT-PCR. Results Fractalkine and CX3CR1 were expressed in the human ovary and in luteinising granulosa cells. However, fractalkine expression was stronger in luteinising granulosa cells. Treatment with fractalkine augmented hCG stimulation of progesterone production in a dose-dependent manner with concomitant increases in transcript levels for key steroidogenic enzymes (StAR, 3-βHSD and CYP11A but had no effect on estradiol biosynthesis(P Conclusions Fractalkine and CX3CR1 were found to express in human ovary and luteinising granulosa cells. Fractalkine can increase the biosynthesis of progesterone in a dose-dependent manner by enhancing transcript levels of key steroidogenic enzymes.

  14. 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.

  15. Homeobox A7 increases cell proliferation by up-regulation of epidermal growth factor receptor expression in human granulosa cells

    Directory of Open Access Journals (Sweden)

    Yanase Toshihiko

    2010-06-01

    Full Text Available Abstract Background Homeobox (HOX genes encode transcription factors, which regulate cell proliferation, differentiation, adhesion, and migration. The deregulation of HOX genes is frequently associated with human reproductive system disorders. However, knowledge regarding the role of HOX genes in human granulosa cells is limited. Methods To determine the role of HOXA7 in the regulation and associated mechanisms of cell proliferation in human granulosa cells, HOXA7 and epidermal growth factor receptor (EGFR expressions were examined in primary granulosa cells (hGCs, an immortalized human granulosa cell line, SVOG, and a granulosa tumor cell line, KGN, by real-time PCR and Western blotting. To manipulate the expression of HOXA7, the HOXA7 specific siRNA was used to knockdown HOXA7 in KGN. Conversely, HOXA7 was overexpressed in SVOG by transfection with the pcDNA3.1-HOAX7 vector. Cell proliferation was measured by the MTT assay. Results Our results show that HOXA7 and EGFR were overexpressed in KGN cells compared to hGCs and SVOG cells. Knockdown of HOXA7 in KGN cells significantly decreased cell proliferation and EGFR expression. Overexpression of HOXA7 in SVOG cells significantly promoted cell growth and EGFR expression. Moreover, the EGF-induced KGN proliferation was abrogated, and the activation of downstream signaling was diminished when HOXA7 was knocked down. Overexpression of HOXA7 in SVOG cells had an opposite effect. Conclusions Our present study reveals a novel mechanistic role for HOXA7 in modulating granulosa cell proliferation via the regulation of EGFR. This finding contributes to the knowledge of the pro-proliferation effect of HOXA7 in granulosa cell growth and differentiation.

  16. Spontaneous transformation of human granulosa cell tumours into an aggressive phenotype: a metastasis model cell line

    International Nuclear Information System (INIS)

    Imai, Misa; Muraki, Miho; Takamatsu, Kiyoshi; Saito, Hidekazu; Seiki, Motoharu; Takahashi, Yuji

    2008-01-01

    Granulosa cell tumours (GCTs) are frequently seen in menopausal women and are relatively indolent. Although the physiological properties of normal granulosa cells have been studied extensively, little is known about the molecular mechanism of GCT progression. Here, we characterise the unique behavioural properties of a granulosa tumour cell line, KGN cells, for the molecular analysis of GCT progression. Population doubling was carried out to examine the proliferation capacity of KGN cells. Moreover, the invasive capacity of these cells was determined using the in vitro invasion assay. The expression level of tumour markers in KGN cells at different passages was then determined by Western blot analysis. Finally, the growth and metastasis of KGN cells injected subcutaneously (s.c.) into nude mice was observed 3 months after injection. During in vitro culture, the advanced passage KGN cells grew 2-fold faster than the early passage cells, as determined by the population doubling assay. Moreover, we found that the advanced passage cells were 2-fold more invasive than the early passage cells. The expression pattern of tumour markers, such as p53, osteopontin, BAX and BAG-1, supported the notion that with passage, KGN cells became more aggressive. Strikingly, KGN cells at both early and advanced passages metastasized to the bowel when injected s.c. into nude mice. In addition, more tumour nodules were formed when the advanced passage cells were implanted. KGN cells cultured in vitro acquire an aggressive phenotype, which was confirmed by the analysis of cellular activities and the expression of biomarkers. Interestingly, KGN cells injected s.c. are metastatic with nodule formation occurring mostly in the bowel. Thus, this cell line is a good model for analysing GCT progression and the mechanism of metastasis in vivo

  17. FOXL2-induced follistatin attenuates activin A-stimulated cell proliferation in human granulosa cell tumors

    Energy Technology Data Exchange (ETDEWEB)

    Cheng, Jung-Chien; Chang, Hsun-Ming; Qiu, Xin; Fang, Lanlan; Leung, Peter C.K., E-mail: peter.leung@ubc.ca

    2014-01-10

    Highlights: •Activin A stimulates cell proliferation in KGN human granulosa cell tumor-derived cell line. •Cyclin D2 mediates activin A-induced KGN cell proliferation. •FOXL2 induces follistatin expression in KGN cells. •FOXL2-induced follistatin attenuates activin A-stimulated KGN cell proliferation. -- Abstract: Human granulosa cell tumors (GCTs) are rare, and their etiology remains largely unknown. Recently, the FOXL2 402C > G (C134W) mutation was found to be specifically expressed in human adult-type GCTs; however, its function in the development of human GCTs is not fully understood. Activins are members of the transforming growth factor-beta superfamily, which has been shown to stimulate normal granulosa cell proliferation; however, little is known regarding the function of activins in human GCTs. In this study, we examined the effect of activin A on cell proliferation in the human GCT-derived cell line KGN. We show that activin A treatment stimulates KGN cell proliferation. Treatment with the activin type I receptor inhibitor SB431542 blocks activin A-stimulated cell proliferation. In addition, our results show that cyclin D2 is induced by treatment with activin A and is involved in activin A-stimulated cell proliferation. Moreover, the activation of Smad signaling is required for activin A-induced cyclin D2 expression. Finally, we show that the overexpression of the wild-type FOXL2 but not the C134W mutant FOXL2 induced follistatin production. Treatment with exogenous follistatin blocks activin A-stimulated cell proliferation, and the overexpression of wild-type FOXL2 attenuates activin A-stimulated cell proliferation. These results suggest that FOXL2 may act as a tumor suppressor in human adult-type GCTs by inducing follistatin expression, which subsequently inhibits activin-stimulated cell proliferation.

  18. Gene expression profiling of upregulated mRNAs in granulosa cells of bovine ovulatory follicles following stimulation with hCG

    Directory of Open Access Journals (Sweden)

    Jacques G. Lussier

    2017-11-01

    Full Text Available Abstract Background Ovulation and luteinization of follicles are complex biological processes initiated by the preovulatory luteinizing hormone surge. The objective of this study was to identify genes that are differentially expressed in bovine granulosa cells (GC of ovulatory follicles. Methods Granulosa cells were collected during the first follicular wave of the bovine estrous cycle from dominant follicles (DF and from ovulatory follicles (OF obtained 24 h following injection of human chorionic gonadotropin (hCG. A granulosa cell subtracted cDNA library (OF-DF was generated using suppression subtractive hybridization and screened. Results Detection of genes known to be upregulated in bovine GC during ovulation, such as ADAMTS1, CAV1, EGR1, MMP1, PLAT, PLA2G4A, PTGES, PTGS2, RGS2, TIMP1, TNFAIP6 and VNN2 validated the physiological model and analytical techniques used. For a subset of genes that were identified for the first time, gene expression profiles were further compared by semiquantitative RT-PCR in follicles obtained at different developmental stages. Results confirmed an induction or upregulation of the respective mRNAs in GC of OF 24 h after hCG-injection compared with those of DF for the following genes: ADAMTS9, ARAF, CAPN2, CRISPLD2, FKBP5, GFPT2, KIT, KITLG, L3MBLT3, MRO, NUDT10, NUDT11, P4HA3, POSTN, PSAP, RBP1, SAT1, SDC4, TIMP2, TNC and USP53. In bovine GC, CRISPLD2 and POSTN mRNA were found as full-length transcript whereas L3MBLT3 mRNA was alternatively spliced resulting in a truncated protein missing the carboxy-terminal end amino acids, 774KNSHNEL780. Conversely, L3MBLT3 is expressed as a full-length mRNA in a bovine endometrial cell line. The 774KNSHNEL780 sequence is well conserved in all mammalian species and follows a SAM domain known to confer protein/protein interactions, which suggest a key function for these amino acids in the epigenetic control of gene expression. Conclusions We conclude that we have identified

  19. Canine ovarian neoplasms: a clinicopathologic study of 71 cases, including histology of 12 granulosa cell tumors.

    Science.gov (United States)

    Patnaik, A K; Greenlee, P G

    1987-11-01

    In a retrospective study of 71 primary ovarian tumors in the dog, epithelial tumors (46%) were more common than sex cord stromal (34%) and germ cell tumors (20%). There were more adenocarcinomas (64%) than adenomas. Sex cord stromal tumors were equally divided into Sertoli-Leydig (12/24) and granulosa cell tumors (12/24). There were equal numbers (7/14) of dysgerminomas and teratomas among the germ cell tumors. Most teratomas (6/7) were malignant. Most granulosa cell tumors were solid; two were mostly cystic. Patterns included sheets of round and ovoid to spindle-shaped cells separated by thin, fibrovascular stroma; neoplastic cells formed rosettes or Call-Exner bodies. In some areas, neoplastic cells were in cords or columns and formed cyst-like structures. Four granulosa cell tumors were macrofollicular, having cysts lined with granulosa cells. Median ages of dogs with different ovarian neoplasms were similar; all were more than 10 years old, except the dogs with teratoma (mean age, 4 years). Most neoplasms were unilateral (84%), except the Sertoli-Leydig cell tumors, many of which were bilateral (36%). Size of ovarian neoplasms varied (2 cm3 to 15,000 cm3). Twenty-nine percent of neoplasms metastasized; adenocarcinomas (48%) and malignant teratomas (50%) had the highest rates, and distant metastasis was more common in malignant teratoma. Endometrial hyperplasia was in 67% of the dogs; it was most common in dogs with sex cord stromal tumors (95%). Uterine malignancy was not seen in dogs with granulosa cell tumors, although hyperplasia endometrium was in all dogs with this tumor. Cysts in the contralateral ovaries were most common in dogs with sex cord stromal tumors.

  20. Androgen and FSH synergistically stimulate lipoprotein degradation and utilization by ovary granulosa cells

    International Nuclear Information System (INIS)

    Schreiber, J.R.; Nakamura, K.; Schmit, V.; Weinstein, D.B.

    1984-01-01

    Androgen can directly modulate the induction of steroidogenic enzymes by FSH (follicle stimulating hormone) in ovary granulosa cells. In studies of its mechanism of action, the authors examined the androgen effect on granulosa cell interaction with lipoproteins, the physiologic source of cholesterol. After granulosa cells were cultured for 48 hours with and without androgen and/or FSH, the cells were incubated for 24 hours with 125 I-lipoproteins [human high density lipoprotein (HDL), rat HDL, or human low density lipoprotein (LDL)]. The media were then analyzed for lipoprotein protein coat degradation products (mainly 125 I-monoiodotyrosine) and progestin [mainly 20 alpha-dihydroprogesterone (20 alpha-DHP)]. In the absence of FSH and androgen, 2 X 10(5) granulosa cells degraded basal levels of all three lipoproteins, but produced no measurable 20 alpha-DHP. The addition of 10(-7) M androstenedione (A), testosterone (T), or 5 alpha-dihydrotestosterone (DHT) had no effect on lipoprotein protein degradation or 20 alpha-DHP production. FSH alone stimulated lipoprotein protein degradation by 50 to 300% while the addition of androgen synergistically augmented the FSH-stimulated 20 alpha-DHP production as well as protein coat degradation of all three lipoproteins. DHT and T were both effective, indicating that androgens themselves, and not estrogen products, were responsible for the effect on lipoprotein protein degradation and 20 alpha-DHP production

  1. Incomplete ovariosalpingectomy and subsequent malignant granulosa cell tumor in a female green iguana (Iguana iguana).

    Science.gov (United States)

    Cruz Cardona, Janice A; Conley, Kenneth J; Wellehan, James F X; Farina, Lisa L; Origgi, Francesco C; Wamsley, Heather L

    2011-07-15

    A 9-year-old spayed female green iguana (Iguana iguana) was evaluated because of a distended coelom and weight loss. History included a single episode of egg binding and subsequent bilateral ovariosalpingectomy. Physical examination revealed a mass within the coelomic cavity. Ultrasonography revealed a large, irregular mass with hypoechoic regions and coelomic effusion. Clinicopathologic derangements included heterophilia, monocytosis, lymphopenia, basophilia, hypocholesterolemia, hypoproteinemia, and hypercalcemia. Results of cytologic evaluation of the mass were suggestive of malignant epithelial neoplasia, but neoplastic cells were not found in the effusion. An ovarian tumor was suspected on the basis of clinical signs, clinicopathologic findings, and results of cytologic evaluation of the mass. Surgical exploration revealed a large left ovary, a normal-appearing contralateral ovary, and a mass in the fat body, all of which were removed and submitted for histologic examination. The histologic diagnosis was granulosa cell tumor with metastasis to the fat body. The patient died 11 months after evaluation, and disseminated granulosa cell tumor was confirmed at necropsy; histologic examination at that time also identified systemic mastocytosis. Granulosa cell tumors are uncommon in reptiles, and this was the first granulosa cell tumor described antemortem cytologically, histologically, and ultrastructurally in an iguana. Findings in this iguana underscored concerns associated with incomplete oophorectomy of iguanas; cytologic and histopathologic findings were similar to those observed in other domestic animals. Oophorectomy should be considered as an alternative to standard ovariosalpingectomy to avoid potential complications in pet reptiles, and use of microsurgical instruments and vascular clips is advised.

  2. Progesterone production requires activation of caspase-3 in preovulatory granulosa cells in a serum starvation model.

    Science.gov (United States)

    An, Li-Sha; Yuan, Xiao-Hua; Hu, Ying; Shi, Zi-Yun; Liu, Xiao-Qin; Qin, Li; Wu, Gui-Qing; Han, Wei; Wang, Ya-Qin; Ma, Xu

    2012-11-01

    Granulosa cells proliferate, differentiate, and undergo apoptosis throughout follicular development. Previous studies have demonstrated that stimulation of progesterone production is accompanied by caspase-3 activation. Moreover, we previously reported that arsenic enhanced caspase-3 activity coupled with progesterone production. Inhibition of caspase-3 activity can significantly inhibit progesterone production induced by arsenic or follicle-stimulating hormone (FSH). Here, we report that serum starvation induces caspase-3 activation coupled with augmentation of progesterone production. Serum starvation also increased the levels of cytochrome P450 cholesterol side chain cleavage enzyme (P450scc) and steroidogenic acute regulatory (StAR) protein, both of which may contribute to progesterone synthesis in preovulatory granulosa cells. Inhibition of caspase-3 activity resulted in a decrease in progesterone production. Deactivation of caspase-3 activity by caspase-3 specific inhibitor also resulted in decreases in P450scc and StAR expression, which may partly contribute to the observed decrease in progesterone production. Our study demonstrates for the first time that progesterone production in preovulatory granulosa cells is required for caspase-3 activation in a serum starvation model. Inhibition of caspase-3 activity can result in decreased expression of the steroidogenic proteins P450scc and StAR. Our work provides further details on the relationship between caspase-3 activation and steroidogenesis and indicates that caspase-3 plays a critical role in progesterone production by granulosa cells. Copyright © 2012 Elsevier Inc. All rights reserved.

  3. 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

    Context:Human granulosa cells (GC) acquire LH receptor (LHR) expression during the follicular phase of the menstrual cycle. Currently, the precise follicular stage is unknown, and specific roles of LH in the follicular development are not fully understood.Objective:Our objective was to measure LH...

  4. Effectiveness of chemotherapy in measurable granulosa cell tumors: a retrospective study and review of literature

    NARCIS (Netherlands)

    van Meurs, Hannah S.; Buist, Marrije R.; Westermann, Anneke M.; Sonke, Gabe S.; Kenter, Gemma G.; van der Velden, Jacobus

    2014-01-01

    Patients with irresectable granulosa cell tumors (GCTs) often receive chemotherapy. The effectiveness of this approach, however, is uncertain. The aim of our study was to assess the response rate to chemotherapy for residual and recurrent inoperable GCT. All consecutive chemotherapy-naive patients

  5. Identification of potential biomarkers in donor cows for in vitro embryo production by granulosa cell transcriptomics

    DEFF Research Database (Denmark)

    Mazzoni, Gianluca; Salleh, Suraya M; Freude, Kristine

    2017-01-01

    The Ovum Pick Up-In vitro Production (OPU-IVP) of embryos is an advanced reproductive technology used in cattle production but the complex biological mechanisms behind IVP outcomes are not fully understood. In this study we sequenced RNA of granulosa cells collected from Holstein cows at oocyte...

  6. Flow cytometric analysis of FSHR, BMRR1B, LHR and apoptosis in granulosa cells and ovulation rate in merino sheep.

    Science.gov (United States)

    Regan, Sheena L P; McFarlane, James R; O'Shea, Tim; Andronicos, Nicholas; Arfuso, Frank; Dharmarajan, Arun; Almahbobi, Ghanim

    2015-08-01

    The aim of the present study was to determine the direct cause of the mutation-induced, increased ovulation rate in Booroola Merino (BB) sheep. Granulosa cells were removed from antral follicles before ovulation and post-ovulation from BB (n=5) and WT (n=12) Merino ewes. Direct immunofluorescence measurement of mature cell surface receptors using flow cytometry demonstrated a significant up-regulation of FSH receptor (FSHR), transforming growth factor beta type 1, bone morphogenetic protein receptor (BMPR1B), and LH receptor (LHR) in BB sheep. The increased density of FSHR and LHR provide novel evidence of a mechanism for increasing the number of follicles that are recruited during dominant follicle selection. The compounding increase in receptors with increasing follicle size maintained the multiple follicles and reduced the apoptosis, which contributed to a high ovulation rate in BB sheep. In addition, we report a mutation-independent mechanism of down-regulation to reduce receptor density of the leading dominant follicle in sheep. The suppression of receptor density coincides with the cessation of mitogenic growth and steroidogenic differentiation as part of the luteinization of the follicle. The BB mutation-induced attenuation of BMPR1B signaling led to an increased density of the FSHR and LHR and a concurrent reduction in apoptosis to increase the ovulation rate. The role of BMPs in receptor modulation is implicated in the development of multiple ovulations. © 2015 Society for Reproduction and Fertility.

  7. Bovine cumulus-granulosa cells contain biologically active retinoid receptors that can respond to retinoic acid

    Directory of Open Access Journals (Sweden)

    Malayer Jerry

    2003-11-01

    Full Text Available Abstract Retinoids, a class of compounds that include retinol and its metabolite, retinoic acid, are absolutely essential for ovarian steroid production, oocyte maturation, and early embryogenesis. Previous studies have detected high concentrations of retinol in bovine large follicles. Further, administration of retinol in vivo and supplementation of retinoic acid during in vitro maturation results in enhanced embryonic development. In the present study, we hypothesized that retinoids administered either in vivo previously or in vitro can exert receptor-mediated effects in cumulus-granulosa cells. Total RNA extracted from in vitro cultured cumulus-granulosa cells was subjected to reverse transcription polymerase chain reaction (RT-PCR and mRNA expression for retinol binding protein (RBP, retinoic acid receptor alpha (RARalpha, retinoic acid receptor beta (RARbeta, retinoic acid receptor gamma (RARgamma, retinoid X receptor alpha (RXRalpha, retinoid X receptor beta (RXRbeta, retinaldehyde dehydrogenase-2 (RALDH-2, and peroxisome proliferator activated receptor gamma (PPARgamma. Transcripts were detected for RBP, RARalpha, RARgamma, RXRalpha, RXRbeta, RALDH-2, and PPARgamma. Expression of RARbeta was not detected in cumulus-granulosa cells. Using western blotting, immunoreactive RARalpha, and RXRbeta protein was also detected in bovine cumulus-granulosa cells. The biological activity of these endogenous retinoid receptors was tested using a transient reporter assay using the pAAV-MCS-betaRARE-Luc vector. Addition of 0.5 and 1 micro molar all-trans retinoic acid significantly (P trans retinol stimulated a mild increase in reporter activity, however, the increase was not statistically significant. Based on these results we conclude that cumulus cells contain endogenously active retinoid receptors and may also be competent to synthesize retinoic acid using the precursor, retinol. These results also indirectly provide evidence that retinoids

  8. Differentiation of Mouse Ovarian Stem Cells Toward Oocyte-Like Structure by Coculture with Granulosa Cells.

    Science.gov (United States)

    Parvari, Soraya; Yazdekhasti, Hossein; Rajabi, Zahra; Gerayeli Malek, Valliollah; Rastegar, Tayebeh; Abbasi, Mehdi

    2016-11-01

    An increasing body of evidence has confirmed existence and function of ovarian stem cells (OSCs). In this study, a novel approach on differentiation of OSCs into oocyte-like cells (OLCs) has been addressed. Recently, different methods have been recruited to isolate and describe aspects of OSCs, but newer and more convenient strategies in isolation are still growing. Herein, a morphology-based method was used to isolate OSCs. Cell suspension of mouse neonatal ovaries was cultured and formed colonies were harvested mechanically and cultivated on mouse embryonic fibroblasts. For differentiation induction, colonies transferred on inactive granulosa cells. Results showed that cells in colonies were positive for alkaline phosphatase activity and reverse transcription-polymerase chain reaction (RT-PCR) confirmed the pluripotency characteristics of cells. Immunofluorescence revealed a positive signal for OCT4, DAZL, MVH, and SSEA1 in colonies as well. Results of RT-PCR and immunofluorescence confirmed that some OLCs were generated within the germ stem cell (GSCs) colonies. The applicability of morphological selection for isolation of GSCs was verified. This method is easier and more economic than other techniques. Our results demonstrate that granulosa cells were effective in inducing the differentiation of OSCs into OLCs through direct cell-to-cell contacts.

  9. Comprehensive analysis of genome-wide DNA methylation across human polycystic ovary syndrome ovary granulosa cell.

    Science.gov (United States)

    Xu, Jiawei; Bao, Xiao; Peng, Zhaofeng; Wang, Linlin; Du, Linqing; Niu, Wenbin; Sun, Yingpu

    2016-05-10

    Polycystic ovary syndrome (PCOS) affects approximately 7% of the reproductive-age women. A growing body of evidence indicated that epigenetic mechanisms contributed to the development of PCOS. The role of DNA modification in human PCOS ovary granulosa cell is still unknown in PCOS progression. Global DNA methylation and hydroxymethylation were detected between PCOS' and controls' granulosa cell. Genome-wide DNA methylation was profiled to investigate the putative function of DNA methylaiton. Selected genes expressions were analyzed between PCOS' and controls' granulosa cell. Our results showed that the granulosa cell global DNA methylation of PCOS patients was significant higher than the controls'. The global DNA hydroxymethylation showed low level and no statistical difference between PCOS and control. 6936 differentially methylated CpG sites were identified between control and PCOS-obesity. 12245 differential methylated CpG sites were detected between control and PCOS-nonobesity group. 5202 methylated CpG sites were significantly differential between PCOS-obesity and PCOS-nonobesity group. Our results showed that DNA methylation not hydroxymethylation altered genome-wide in PCOS granulosa cell. The different methylation genes were enriched in development protein, transcription factor activity, alternative splicing, sequence-specific DNA binding and embryonic morphogenesis. YWHAQ, NCF2, DHRS9 and SCNA were up-regulation in PCOS-obesity patients with no significance different between control and PCOS-nonobesity patients, which may be activated by lower DNA methylaiton. Global and genome-wide DNA methylation alteration may contribute to different genes expression and PCOS clinical pathology.

  10. Lower FOXO3 mRNA expression in granulosa cells is involved in unexplained infertility.

    Science.gov (United States)

    Yamamoto, Hikaru; Yamashita, Yoshiki; Saito, Natsuho; Hayashi, Atsushi; Hayashi, Masami; Terai, Yoshito; Ohmichi, Masahide

    2017-06-01

    The aim of this study was to investigate whether FOXO1 and FOXO3 mRNA expression in granulosa cells is the cause of unexplained infertility. Thirty-one patients aged infertility and 18 with male partner infertility as a control group) whose serum anti-Müllerian hormone level was >0.5 ng/μL were enrolled in the study. All patients underwent oocyte retrieval under a short protocol from June 2012 to October 2013. Real-time PCR was carried out using mRNA extracted from granulosa cells retrieved from mature follicles. We compared FOXO1 and FOXO3 mRNA expression ratios in granulosa cells between the unexplained infertility group and the male infertility group. The relation between FOXO1 and FOXO3 mRNA expression ratios in granulosa cells and assisted reproduction technology clinical outcome was also examined. FOXO3 mRNA expression ratio was significantly lower in the unexplained infertility group than in the male infertility group. Moreover, FOXO3 mRNA expression ratio showed a positive correlation with both the number of retrieved oocytes and serum anti-Müllerian hormone level. A positive correlation was also identified between FOXO1 mRNA expression and total dose of hMG. As well, the number of retrieved oocytes in the unexplained infertility group was statistically lower than that in the male infertility group. A lower FOXO3 mRNA expression in granulosa cells leads to poor oocyte development in patients with unexplained infertility undergoing controlled ovarian stimulation for in vitro fertilization-embryo transfer. © 2017 Japan Society of Obstetrics and Gynecology.

  11. Evidence for ovarian granulosa stem cells: telomerase activity and localization of the telomerase ribonucleic acid component in bovine ovarian follicles.

    Science.gov (United States)

    Lavranos, T C; Mathis, J M; Latham, S E; Kalionis, B; Shay, J W; Rodgers, R J

    1999-08-01

    We have previously postulated that granulosa cells of developing follicles arise from a population of stem cells. Stem cells and cancer cells can divide indefinitely partly because they express telomerase. Telomerase is a ribonucleoprotein enzyme that repairs the ends of telomeres that otherwise shorten progressively upon each successive cell division. In this study we carried out cell cycle analyses and examined telomerase expression to examine our hypothesis. Preantral (60-100 microm) and small (1 mm) follicles, as well as granulosa cells from medium-sized (3 mm) and large (6-8 mm) follicles, were isolated. Cell cycle analyses and expression of Ki-67, a cell cycle-related protein, were undertaken on follicles of each size (n = 3) by flow cytometry; 12% to 16% of granulosa cells in all follicles were in the S phase, and less than 2% were in the G(2)/M phase. Telomerase activity (n = 3) was highest in the small preantral follicles, declining at the 1-mm stage and even further at the 3-mm stage. In situ hybridization histochemistry was carried out on bovine ovaries, and telomerase RNA was detected in the granulosa cells of growing follicles but not primordial follicles. Two major patterns of staining were observed in the membrana granulosa of antral follicles: staining in the middle and antral layers, and staining in the middle and basal layers. No staining was detected in oocytes. Our results strongly support our hypothesis that granulosa cells arise from a population of stem cells.

  12. Transient expression of progesterone receptor and cathepsin-l in human granulosa cells during the periovulatory period.

    Science.gov (United States)

    García, Víctor; Kohen, Paulina; Maldonado, Carola; Sierralta, Walter; Muñoz, Alex; Villarroel, Claudio; Strauss, Jerome F; Devoto, Luigi

    2012-03-01

    To study in vivo the progesterone receptor (PR) expression levels in human granulosa cells (GCs) during the periovulatory period and the affect of the protein kinase A (PKA) pathway on PR expression and cathepsin-L expression-activation. Experimental study. University research unit. Twenty-five women of reproductive age. Follicular fluid and GCs obtained from spontaneous cycles before and during the normal luteinizing hormone surge, and samples obtained 36 hours after human chorionic gonadotropin (hCG) administration in patients undergoing in vitro fertilization. To determine PR, cathepsin-L messenger RNA (mRNA) analysis via real-time polymerase chain reaction, and protein of PR, cathepsin-L, and PKA in human GCs. The Western blot analysis revealed that bands of PR (isoform A) were the most abundant and that mRNA (PR-A and PR-B) have a temporal pattern of expression throughout the periovulatory period. The protein levels of PR and cathepsin-L were up-regulated by hCG. The abundance of PR was diminished in the presence of PKA inhibitor, and cathepsin-L with PR receptor antagonist. The transient expression of PR in human GCs of the preovulatory follicle suggests that PR and its ligand play a role in the activation of cathepsin-L, which is presumably involved in the degradation of the follicular extracellular matrix during human ovulation. Copyright © 2012 American Society for Reproductive Medicine. All rights reserved.

  13. Wilms' Tumor 1 Overexpression in Granulosa Cells Is Associated with Polycystic Ovaries in Polycystic Ovary Syndrome Patients.

    Science.gov (United States)

    Wang, Qun; Huang, Tao; Shu, Xin; Zhao, Shi-Gang; Liang, Yu; Muhammad, Tahir; Gao, Fei; Zhao, Han; Liu, Hong-Bin

    2018-01-01

    Polycystic ovary syndrome (PCOS) is a heterogeneous disorder characterized by chronic ovulatory dysfunction, hyperandrogenism, and polycystic ovaries. Wilms' tumor 1 (WT1) encoding a transcription factor involved in the differentiation of granulosa cells (GCs) regulates androgen receptor in the development of male genitalia. However, the expression pattern and possible role of WT1 in ovaries of PCOS patients are still unknown. GCs from 95 PCOS patients (PCOS group) and 62 healthy controls (control group) were isolated. The expression of WT1 in GCs was quantified using the reverse transcription-polymerase chain reaction. The correlation between WT1 expression and clinical characteristics was evaluated in PCOS patients. WT1 expression was increased in PCOS patients compared with the normal controls. The expression of WT1 was moderately correlated with testosterone (r = 0.334, p = 0.001) and luteinizing hormone (r = 0.357, p = 0.001) levels and the antral follicle counts (r = 0.337, p = 0.001). Our study provided novel insights into the relationship between hyperandrogenism and polycystic ovaries of PCOS and WT1. © 2018 S. Karger AG, Basel.

  14. Detection of calmodulin binding protein at 170 KDA in BALB, AKR, DON and chicken granulosa cells

    International Nuclear Information System (INIS)

    Selinfreund, R.; Lin, P.H.; Marrone, B.; Wharton, W.

    1987-01-01

    Calmodulin (CAM) has been shown to bind to the epidermal growth factor (EGF) receptor (170 kDa) and is phosphorylated in a EGF dependent manner in the A431 human epidermoid carcinoma cells. In the present study, they report 125 I-CAM binding to a 170 kDa protein detected in cell membrane vesicles of Balb/3T3, AKR, DON and chicken granulosa cells. Purified plasma membranes from these cells were resolved via electrophoresis (without heat denaturation) and electroblotted onto nictrocellulose paper. Upon hybridizing against 125 I-CAM, a distinct autoradiographic band occurred at 170 kDa for all the cells lines under study. The binding of CAM is specific and can be displaced with the addition of excess unlabeled CAM. The result suggest that 125 I-CAM may bind to the 170 kDa EGF receptor in BALB, AKR, DON and chicken granulosa cells

  15. Effect of anabolics on bovine granulosa-luteal cell primary cultures.

    Directory of Open Access Journals (Sweden)

    Bartolomeo Biolatti

    2007-10-01

    Full Text Available Granulosa cell tumours are observed with increased frequency among calves slaughtered in Northern Italy. The use of illegal anabolics in breeding was taken into account as a cause of this pathology. An in vitro approach was used to detect the possible alterations of cell proliferation induced by anabolics on primary cultures of bovine granulosa-luteal cells. Cultures were treated with different concentrations of substances illegally used in cattle (17beta-estradiol, clenbuterol and boldione. Cytotoxicity was determined by means of MTT test, to exclude toxic effects induced by anabolics and to determine the highest concentration to be tested. Morphological changes were evaluated by means of routine cytology, while PCNA expression was quantified in order to estimate cell proliferation. Cytotoxic effects were revealed at the highest concentrations. The only stimulating effect on cell proliferation was detected in boldione treated cultures: after 48 h treated cells, compared to controls, showed a doubled expression of PCNA. In clenbuterol and 17beta-estradiol treated cells PCNA expression was similar to controls or even decreased. As the data suggest an alteration in cell proliferation, boldione could have a role in the early stage of pathogenesis of granulosa cell tumour in cattle.

  16. Docosahexaenoic acid (DHA) effects on proliferation and steroidogenesis of bovine granulosa cells.

    Science.gov (United States)

    Maillard, Virginie; Desmarchais, Alice; Durcin, Maeva; Uzbekova, Svetlana; Elis, Sebastien

    2018-04-26

    Docosahexaenoic acid (DHA) is a n-3 polyunsaturated fatty acid (PUFA) belonging to a family of biologically active fatty acids (FA), which are known to have numerous health benefits. N-3 PUFAs affect reproduction in cattle, and notably directly affect follicular cells. In terms of reproduction in cattle, n-3 PUFA-enriched diets lead to increased follicle size or numbers. The objective of the present study was to analyze the effects of DHA (1, 10, 20 and 50 μM) on proliferation and steroidogenesis (parametric and/or non parametric (permutational) ANOVA) of bovine granulosa cells in vitro and mechanisms of action through protein expression (Kruskal-Wallis) and signaling pathways (non parametric ANOVA) and to investigate whether DHA could exert part of its action through the free fatty acid receptor 4 (FFAR4). DHA (10 and 50 μM) increased granulosa cell proliferation and DHA 10 μM led to a corresponding increase in proliferating cell nuclear antigen (PCNA) expression level. DHA also increased progesterone secretion at 1, 20 and 50 μM, and estradiol secretion at 1, 10 and 20 μM. Consistent increases in protein levels were also reported for the steroidogenic enzymes, cytochrome P450 family 11 subfamily A member 1 (CYP11A1) and hydroxy-delta-5-steroid dehydrogenase, 3 beta- and steroid delta-isomerase 1 (HSD3B1), and of the cholesterol transporter steroidogenic acute regulatory protein (StAR), which are necessary for production of progesterone or androstenedione. FFAR4 was expressed in all cellular types of bovine ovarian follicles, and in granulosa cells it was localized close to the cellular membrane. TUG-891 treatment (1 and 50 μM), a FFAR4 agonist, increased granulosa cell proliferation and MAPK14 phosphorylation in a similar way to that observed with DHA treatment. However, TUG-891 treatment (1, 10 and 50 μM) showed no effect on progesterone or estradiol secretion. These data show that DHA stimulated proliferation and steroidogenesis of bovine

  17. Adult type granulosa cell tumor in adult testis: report of a case and review of the literature

    Directory of Open Access Journals (Sweden)

    Zhanyong Bing

    2011-10-01

    Full Text Available Granulosa cell tumors can be classified into juvenile and adult types and more commonly occur in ovaries. Adult testicular granulosa cell tumors are extremely rare and only 29 cases of adult type have previously been reported. We report here a 28-year-old Caucasian man with a left testicular adult type granulosa cell tumor. The tumor measured 2.6 x 2.6 x 2.5 cm and was mitotically active (10/10 HPF. Immunohistochemical stains showed the tumor diffusely positive for inhibin and vimentin, and negative for epithelial membrane antigen, cytokeratins, synaptophysin, HMB-45, OCT-4, placental-like alkaline phosphatase and lymphoid markers . The reported granulosa cell tumors in adult testis were briefly reviewed.

  18. 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.

  19. PMS2 gene mutation results in DNA mismatch repair system failure in a case of adult granulosa cell tumor.

    Science.gov (United States)

    Wang, Wen-Chung; Lee, Ya-Ting; Lai, Yen-Chein

    2017-03-27

    Granulosa cell tumors are rare ovarian malignancies. Their characteristics include unpredictable indolent growth with malignant potential and late recurrence. Approximately 95% are of adult type. Recent molecular studies have characterized the FOXL2 402C > G mutation in adult granulosa cell tumor. Our previous case report showed that unique FOXL2 402C > G mutation and defective DNA mismatch repair system are associated with the development of adult granulosa cell tumor. In this study, the DNA sequences of four genes, MSH2, MLH1, MSH6, and PMS2, in the DNA mismatch repair system were determined via direct sequencing to elucidate the exact mechanism for the development of this granulosa cell tumor. The results showed that two missense germline mutations, T485K and N775L, inactivate the PMS2 gene. The results of this case study indicated that although FOXL2 402C > G mutation determines the development of granulosa cell tumor, PMS2 mutation may be the initial driver of carcinogenesis. Immunohistochemistry-based tumor testing for mismatch repair gene expression may be necessary for granulosa cell tumors to determine their malignant potential or if they are part of Lynch syndrome.

  20. 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

    2017-10-01

    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.

  1. A Low-Testosterone State Associated with Endometrioma Leads to the Apoptosis of Granulosa Cells

    Science.gov (United States)

    Ono, Yoshihiro J.; Tanabe, Akiko; Nakamura, Yoko; Yamamoto, Hikaru; Hayashi, Atsushi; Tanaka, Tomohito; Sasaki, Hiroshi; Hayashi, Masami; Terai, Yoshito; Ohmichi, Masahide

    2014-01-01

    Although endometriosis is suspected to be a cause of premature ovarian insufficiency (POI), the mechanism(s) underlying this process have not been elucidated. Recently, androgens were shown to promote oocyte maturation and to play a role in folliculogenesis. In addition, several reports have documented low testosterone levels in the follicular fluid obtained from endometriosis patients. We therefore examined whether the low levels of serum testosterone are associated with the apoptosis of granulosa cells in follicles obtained from endometriosis patients. Serum samples were collected from 46 patients with endometriosis and from 62 patients without endometriosis who received assisted reproductive therapy. Specimens of the ovaries obtained from 10 patients with endometrioma were collected using laparoscopy. The mean serum testosterone concentration in the patients with endometriosis was significantly lower than that observed in the patients without endometriosis. Furthermore, high expression of a pro-apoptotic Bcl-2 member, BimEL, in the follicles was found to be associated with a low serum testosterone level. We clarified the underlying mechanisms using a basic approach employing human immortalized granulosa cells derived from a primary human granulosa cell tumor, the COV434 cell line. The in vitro examination demonstrated that testosterone inhibited apoptosis induced by sex steroids depletion via the PI3K/Akt-FoxO3a pathway in the COV434 cells. In conclusion, we elucidated the mechanism underlying the anti-apoptotic effects of testosterone on granulosa cells, and found that a low-testosterone status is a potentially important step in the development of premature ovarian insufficiency in patients with endometriosis. PMID:25536335

  2. A low-testosterone state associated with endometrioma leads to the apoptosis of granulosa cells.

    Directory of Open Access Journals (Sweden)

    Yoshihiro J Ono

    Full Text Available Although endometriosis is suspected to be a cause of premature ovarian insufficiency (POI, the mechanism(s underlying this process have not been elucidated. Recently, androgens were shown to promote oocyte maturation and to play a role in folliculogenesis. In addition, several reports have documented low testosterone levels in the follicular fluid obtained from endometriosis patients. We therefore examined whether the low levels of serum testosterone are associated with the apoptosis of granulosa cells in follicles obtained from endometriosis patients. Serum samples were collected from 46 patients with endometriosis and from 62 patients without endometriosis who received assisted reproductive therapy. Specimens of the ovaries obtained from 10 patients with endometrioma were collected using laparoscopy. The mean serum testosterone concentration in the patients with endometriosis was significantly lower than that observed in the patients without endometriosis. Furthermore, high expression of a pro-apoptotic Bcl-2 member, BimEL, in the follicles was found to be associated with a low serum testosterone level. We clarified the underlying mechanisms using a basic approach employing human immortalized granulosa cells derived from a primary human granulosa cell tumor, the COV434 cell line. The in vitro examination demonstrated that testosterone inhibited apoptosis induced by sex steroids depletion via the PI3K/Akt-FoxO3a pathway in the COV434 cells. In conclusion, we elucidated the mechanism underlying the anti-apoptotic effects of testosterone on granulosa cells, and found that a low-testosterone status is a potentially important step in the development of premature ovarian insufficiency in patients with endometriosis.

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

    Directory of Open Access Journals (Sweden)

    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.

  4. Human luteinized granulosa cells secrete apoB100-containing lipoproteins

    NARCIS (Netherlands)

    Gautier, Thomas; Becker, Steffi; Drouineaud, Veronique; Menetrier, Franck; Sagot, Paul; Nofer, Jerzy-Roch; von Otte, Soeren; Lagrost, Laurent; Masson, David; Tietge, Uwe J. F.

    Thus far, liver, intestine, heart, and placenta have been shown to secrete apolipoprotein (apo) B-containing lipoproteins. In the present study, we first investigated lipoproteins in human follicular fluid (FF), surrounding developing oocytes within the ovary, as well as in corresponding plasma

  5. Significantly lengthened telomere in granulosa cells from women with polycystic ovarian syndrome (PCOS).

    Science.gov (United States)

    Wei, Duo; Xie, Juanke; Yin, Baoli; Hao, Haoying; Song, Xiaobing; Liu, Qi; Zhang, Cuilian; Sun, Yingpu

    2017-07-01

    Polycystic ovary syndrome (PCOS) is the most common endocrinopathy among women at reproductive age. However, its etiology remains poorly understood. Recent studies indicated that telomere length was related to PCOS. However, the association between telomere length and PCOS has only been shown in leucocytes and remained controversial across different studies. To clarify the association between telomere length and PCOS, the current study interrogated telomere length not only in leucocytes, but also in follicular granulosa cells, which is essential for folliculogenesis and steroidogenesis. Seventy-five patients with PCOS and 81 controls with mechanical infertility undergoing their first in vitro fertilization cycle were enrolled. Their peripheral blood and granulosa cells were collected on the oocyte retrieval day. Telomere length of both leucocytes in the blood and granulosa cells was assayed by quantitative polymerase chain reaction. No significant difference was found in the leucocyte telomere length between controls and PCOS patients (0.99 ± 0.44 vs. 1.00 ± 0.38, p = 0.93). Interestingly, when comparing telomere length in granulosa cells between controls and PCOS subjects, significantly lengthened telomere length was found in PCOS subjects (1.00 ± 0.37 vs. 1.57±0.67, p < 0.0001). After adjustments for age and body mass index, the p value remained significant (p < 0.0001). This finding reinforced the association between telomere abnormalities and PCOS. Given the importance of telomere length in cellular proliferation, our findings provided novel insights into the pathophysiology of PCOS that abnormalities in telomere length possibly disturb folliculogenesis and subsequently result in PCOS.

  6. 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.

  7. ANTIOXIDANT STATUS AND EXPRESSION OF HEAT SHOCK PROTEIN OF COBALT-TREATED PORCINE OVARIAN GRANULOSA CELLS

    Directory of Open Access Journals (Sweden)

    Marcela Capcarová

    2013-02-01

    Full Text Available The aim of this study was to determine the activity of superoxide dismutase (SOD, total antioxidant status (TAS and expression of heat shock protein 70 (Hsp70 of porcine ovarian granulosa cells cultured in vitro after cobalt (Co administrations. Ovarian granulosa cells were incubated with cobalt sulphate administrations as follows: group E1 (0.09 mg.ml-1, group E2 (0.13 mg.ml-1, group E3 (0.17 mg.ml-1, group E4 (0.33 mg.ml-1, group E5 (0.5 mg.ml-1 and the control group without any additions for 18 h. Co administration developed stress reaction and promoted accumulation of Hsp70 what resulted in increasing activity of SOD. TAS of granulosa cells increased with higher doses of Co whereas low doses had no effect on this parameter. Trace elements can adversely affect animal female reproductive system and its functions, through either direct or indirect effects on oxidative stress induction.

  8. 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.

  9. The anti-epileptic drug valproic acid (VPA inhibits steroidogenesis in bovine theca and granulosa cells in vitro.

    Directory of Open Access Journals (Sweden)

    Claire Glister

    Full Text Available Valproic acid (VPA is used widely to treat epilepsy and bipolar disorder. Women undergoing VPA treatment reportedly have an increased incidence of polycystic ovarian syndrome (PCOS-like symptoms including hyperandrogenism and oligo- or amenorrhoea. To investigate potential direct effects of VPA on ovarian steroidogenesis we used primary bovine theca (TC and granulosa (GC cells maintained under conditions that preserve their 'follicular' phenotype. Effects of VPA (7.8-500 µg/ml on TC were tested with/without LH. Effects of VPA on GC were tested with/without FSH or IGF analogue. VPA reduced (P99% decrease; P<0.0001 with lesser effects on LHR, STAR, CYP11A1 and HSD3B1 mRNA (<90% decrease; P<0.05. VPA only reduced TC progesterone secretion induced by the highest (luteinizing LH dose tested; TC number was unaffected by VPA. At higher concentrations (125-500 µg/ml VPA inhibited basal, FSH- and IGF-stimulated estradiol secretion (P<0.0001 by GC without affecting progesterone secretion or cell number. VPA reversed FSH-induced upregulation of CYP19A1 and HSD17B1 mRNA abundance (P<0.001. The potent histone deacetylase (HDAC inhibitors trichostatin A and scriptaid also suppressed TC androstenedione secretion and granulosal cell oestrogen secretion suggesting that the action of VPA reflects its HDAC inhibitory properties. In conclusion, these findings refute the hypothesis that VPA has a direct stimulatory action on TC androgen output. On the contrary, VPA inhibits both LH-dependent androgen production and FSH/IGF-dependent estradiol production in this in vitro bovine model, likely by inhibition of HDAC.

  10. IL-6 Promotes FSH-Induced VEGF Expression Through JAK/STAT3 Signaling Pathway in Bovine Granulosa Cells

    Directory of Open Access Journals (Sweden)

    Meng Yang

    2017-11-01

    Full Text Available Background/Aims: Vascular endothelial growth factor (VEGF has been demonstrated to play a pivotal role in the regulation of angiogenesis in ovarian follicular development, particularly during the preovulatory period. Although numerous studies have shown that interleukin-6 (IL-6 is one of the major inducing factors that regulate the expression of VEGF in non-ovarian cells, whether it involved in regulating the expression of VEGF in normal ovarian granulosa cells is still unknown. The aim of this study was to elucidate the mechanisms underlying the effect of IL-6 on FSH-induced VEGF expression in bovine granulosa cells derived from large follicles. Methods: VEGF mRNA expression in granulosa cells after IL-6 with/without inhibitors treatment was analyzed by RT-qPCR. Phosphorylation levels of ERK1/2 and STAT3 proteins induced by IL-6 were analyzed by western blotting. The protein levels produced by granulosa cells were detected by ELISA. Results: High concentration of IL-6 (10ng/ml can significantly up-regulate FSH-induced VEGF gene and protein expression levels in granulosa cells, and also promote the VEGF upstream regulators HIF-1α and COX2 mRNA expression. VEGF expression levels were significantly decreased after specifically blocking HIF-1α and COX2 by using inhibitors. The up-regulation effect of IL-6 on FSH-induced VEGF expression in granulosa cells mainly through activating the JAK/STAT3 signaling pathway, which can be impaired by JAK inhibitors. Conclusion: IL-6 can promote FSH-induced VEGF expression in granulosa cells, which is mainly achieved by increasing the expression of HIF-1α and COX2.This promoting effect is mediated by activating the JAK/STAT3 pathway. Moreover, there may be a synergistic relationship between FSH and IL-6 in the regulation of VEGF expression.

  11. 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

  12. 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.

  13. Cancer of rat ovaries: Sertoli cell or granulosa-theca cell tumours

    International Nuclear Information System (INIS)

    Knowles, J.F.

    1983-01-01

    The effects of X-radiation (0-1.25 Gy) given 24 hours after neonatal injections of the carcinogen ethyl nitrosourea (ENU) (0-10 mg/kg) in female rats were studied. Twelve out of 118 rats bore single ovarian tumours. A substantial excess of ovarian tumours occurred in the rats given 4 mg/kg ENU and 1.25 GY X-rays but not in others given ENU alone, radiation alone or 10 mg/kg ENU and 1.25 Gy. The tumours were all found in old rats (657-1085 days). In all of the tumours the presence of tubular formations suggested a diagnosis of ovarian Sertoli cell tumour. In two tumours, only a few tubular structures were seen and fibrous stromal tissue predominated, suggesting a diagnosis of granulosa-theca cell tumour. All other tumours were a mixture of both elements. (U.K.)

  14. Effects of the Macular Carotenoid Lutein in Human Retinal Pigment Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Xiaoming Gong

    2017-12-01

    Full Text Available Retinal pigment epithelial (RPE cells are central to retinal health and homoeostasis. Oxidative stress-induced damage to the RPE occurs as part of the pathogenesis of age-related macular degeneration and neovascular retinopathies (e.g., retinopathy of prematurity, diabetic retinopathy. The xanthophyll carotenoids, lutein and zeaxanthin, are selectively taken up by the RPE, preferentially accumulated in the human macula, and transferred to photoreceptors. These macular xanthophylls protect the macula (and the broader retina via their antioxidant and photo-protective activities. This study was designed to investigate effects of various carotenoids (β-carotene, lycopene, and lutein on RPE cells subjected to either hypoxia or oxidative stress, in order to determine if there is effect specificity for macular pigment carotenoids. Using human RPE-derived ARPE-19 cells as an in vitro model, we exposed RPE cells to various concentrations of the specific carotenoids, followed by either graded hypoxia or oxidative stress using tert-butyl hydroperoxide (tBHP. The results indicate that lutein and lycopene, but not β-carotene, inhibit cell growth in undifferentiated ARPE-19 cells. Moreover, cell viability was decreased under hypoxic conditions. Pre-incubation of ARPE-19 cells with lutein or lycopene protected against tBHP-induced cell loss and cell co-exposure of lutein or lycopene with tBHP essentially neutralized tBHP-dependent cell death at tBHP concentrations up to 500 μM. Our findings indicate that lutein and lycopene inhibit the growth of human RPE cells and protect the RPE against oxidative stress-induced cell loss. These findings contribute to the understanding of the protective mechanisms attributable to retinal xanthophylls in eye health and retinopathies.

  15. Modulation of cultured porcine granulosa cell responsiveness to follicle stimulating hormone and epidermal growth factor

    Energy Technology Data Exchange (ETDEWEB)

    Buck, P.A.

    1986-01-01

    Ovarian follicular development is dependent upon the coordinated growth and differentiation of the granulosa cells which line the follicle. Follicle stimulating hormone (FSH) induces granulosa cell differentiation both in vivo and in vitro. Epidermal growth factor (EGF) stimulates granulosa cell proliferation in vitro. The interaction of these two effectors upon selected parameters of growth and differentiation was examined in monolayer cultures of porcine granulose cells. Analysis of the EGF receptor by /sup 125/I-EGF binding revealed that the receptor was of high affinity with an apparent dissociation constant of 4-6 x 10/sup -10/ M. The average number of receptors per cell varied with the state of differentiation both in vivo and in vitro; highly differentiated cells bound two-fold less /sup 125/I-EGF and this effect was at least partially induced by FSH in vitro. EGF receptor function was examined by assessing EGF effects on cell number and /sup 3/H-thymidine incorporation. EGF stimulated thymidine incorporation in both serum-free and serum-supplemented culture, but only in serum-supplemented conditions was cell number increased. EGF receptor function was inversely related to the state of differentiation and was attenuated by FSH. The FSH receptor was examined by /sup 125/I-FSH binding. EGF increased FSH receptor number, and lowered the affinity of the receptor. The function of these receptors was assessed by /sup 125/I-hCG binding and progesterone radioimmunoassay. If EGF was present continuously in the cultures. FSH receptor function was attenuated regardless of FSH receptor number. A preliminary effort to examine the mechanism of this interaction was performed by analyzing hormonally controlled protein synthesis with /sup 35/S-methionine labeling, SDS polyacrylamide gel electrophoresis and fluorography. FSH promoted the expression of a 27,000 dalton protein. This effect was attenuated by EGF.

  16. TLR4 activates NF-κB in human ovarian granulosa tumor cells

    International Nuclear Information System (INIS)

    Woods, Dori C.; White, Yvonne A.R.; Dau, Caroline; Johnson, A.L.

    2011-01-01

    Highlights: → TLR4 is expressed in human ovarian granulosa tumor cells. → Acting through TLR4, LPS and HSP60 induce a NFκB signaling cascade in human ovarian granulosa tumor cells. → NFκB 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-κB) 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 IκB degradation and activation of NF-κB. NF-κB activation was confirmed by nuclear localization of NF-κB p65 following treatment with LPS and the naturally occurring ligand, HSP60. Notably, immunoneutralization of TLR4 blocked nuclear localization, and inhibition of NF-κB signaling attenuated LPS-induced TNFα plus increased doubling time in both cell lines. Contradictory to reports using human OSE cell lines, inhibition of NF-κB 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-κB does not sensitize GCTs to TRAIL or cisplatin.

  17. Localization and movement of newly synthesized cholesterol in rat ovarian granulosa cells

    International Nuclear Information System (INIS)

    Lange, Y.; Schmit, V.M.; Schreiber, J.R.

    1988-01-01

    The distribution and movement of cholesterol were studied in granulosa cells from the ovaries of estrogen-stimulated hypophysectomized immature rats cultured in serum-free medium. Plasma membrane cholesterol was distinguished from intracellular cholesterol with cholesterol oxidase, an enzyme that converts cell surface cholesterol to cholestenone, leaving intracellular cholesterol untouched. Using this approach we showed that 82% of unesterified cholesterol was associated with the plasma membrane in granulosa cells cultured for 48 h in serum-free medium in both the presence and absence of added androstenedione and FSH. FSH and androstenedione stimulated a marked increase in steroid hormone (progestin) production. The movement of newly synthesized cholesterol to the plasma membrane also was followed using cholesterol oxidase. Newly synthesized cholesterol reached the plasma membrane too rapidly to be measured in unstimulated cells (t1/2 less than 20 min); however, in cells stimulated by FSH and androstenedione, this rate was considerably slower (t1/2 approximately 2h). Therefore, cholesterol movement to the plasma membrane appears to be regulated by gonadotropins in these cells. We tested whether steroid biosynthesis used all cell cholesterol pools equally. To this end we administered [3H]acetate and [14C]acetate at different times and determined their relative specific contents in various steroids after defined intervals. The relative ages of the steroids (youngest to oldest) were: lanosterol, progestins, intracellular cholesterol, and plasma membrane cholesterol. This finding suggests that progestins use newly synthesized intracellular cholesterol in preference to preexisting intracellular or cell surface cholesterol

  18. TGF-β1 resulting in differential microRNA expression in bovine granulosa cells.

    Science.gov (United States)

    Xu, Yefen; Niu, Jiaqiang; Xi, Guangying; Niu, Xuezhi; Wang, Yuheng; Guo, Ming; Yangzong, Qiangba; Yao, Yilong; Sizhu, Suo Lang; Tian, Jianhui

    2018-07-15

    To explore the expression profile of the cellular miRNAs in bovine ovarian granulosa cells responding to transforming growth factor-β1 (TGF-β1), the effect of TGF-β1 on cell proliferation was firstly investigated by CCK-8 method and the results showed that there was a significant inhibitory effect on bovine granulosa cell proliferation treated with 5/10 ng/mL human recombinant TGF-β1 for 24 h compared to the control (P cells stimulated with or without 10 ng/mL human recombinant TGF-β1. A total of 13,257,248 and 138,726,391 clean reads per library were obtained from TGF-β1 and control groups, respectively. There were 498 and 499 bovine-specific exist miRNAs (exist miRNAs), 627 and 570 conserved known miRNAs (known miRNAs), and 593 and 585 predicted novel miRNAs in TGF-β1 and control groups, respectively. A total of 78 miRNAs with significant differential expression, including 39 up-regulated miRNAs and 39 down-regulated miRNAs were identified in the TGF-β1 group compared with the control. Real-time quantitative PCR analyses of bta-miR-106a and bta-miR-1434-5p showed that their up-expressions were interrupted by SB431542, an inhibitor that blocks TGFβ1/Smad signaling, which supported the sequencing data. GO analysis showed involvement of the predicted genes of the differentially expressed miRNAs in a broad spectrum of cell biological processes, cell components, and molecular functions. KEGG pathway analysis of the predicted miRNA targets further indicated that these differentially expressed miRNAs are involved in various signaling pathways, such as Wnt, MAPK, and TGF-β signaling, which might be involved in follicular development. These results provide valuable information on the composition, expression, and function of miRNAs in bovine granulosa cells responding to TGF-β1, and will aid in understanding the molecular mechanisms of TGF-β1 in granulosa cells. Copyright © 2018 Elsevier B.V. All rights reserved.

  19. Functional study and regional mapping of 44 hormono-regulated genes isolated from a porcine granulosa cell library

    Directory of Open Access Journals (Sweden)

    Hatey François

    2001-01-01

    Full Text Available Abstract cDNA clones from a pig granulosa cell cDNA library were isolated by differential hybridisation for follicle stimulating hormone (FSH regulation in granulosa cells in a previous study. The clones that did not match any known sequence were studied for their expression in granulosa cells (treated or not by FSH and in fresh isolated ovarian follicles mainly by comparative RT-PCR analysis. These results give functional data on genes that may be implicated in follicular growing. These ESTs have been localised on the porcine genome, using a somatic cell hybrid panel, providing new type I markers on the porcine map and information on the comparative map between humans and pigs.

  20. EFFECT OF dbcAMP ON PROLIFERATION AND APOPTOSIS OF PORCINE GRANULOSA CELLS in vitro

    Directory of Open Access Journals (Sweden)

    Richard Alexa

    2013-02-01

    Full Text Available Cyclic nucleotide cAMP and its target protein kinase A (PKA dependent intracellular mechanisms can play an important role in regulation of ovarian cell function and in mediating gonadotropin action on these cells. The aim of the present study was to examine the effect of cAMP analogue, dibutyryl cyclic adenosine monophosphate (dbcAMP (0; 0.1; 1 and 10 µg/ml or FSH (0; 0,01; 1 IU/ml on proliferation and apoptosis of porcine granulosa cells in vitro. Indices of cell apoptosis (expression of apoptotic peptide bax and proliferation (expression of proliferation-associated peptide PCNA within ovarian granulosa cells were analysed by immunocytochemistry. It was observed that accumulation of PCNA was increased by dbcAMP and FSH at all doses added. The occurrence of bax was also stimulated by dbcAMP after exposition (at 0,1 and 1 µg/ml, but not at dose 10 µg/ml and by FSH (at all doses added. The stimulatory effect of both dbcAMP and FSH on both ovarian cell apoptosis and proliferation suggest, that these substances may promote ovarian follicular cell turnover. The similarity of dbcAMP and FSH effect may indicate that FSH can affect ovarian functions via cAMP-dependent intracellular mechanisms. The present data may provide new tools to regulate human and animal reproductive processes via cAMP-dependent mechanisms.

  1. In vitro culture of oocytes and granulosa cells collected from normal, obese, emaciated and metabolically stressed ewes.

    Science.gov (United States)

    Tripathi, S K; Farman, M; Nandi, S; Mondal, S; Gupta, Psp; Kumar, V Girish

    2016-07-01

    The present study was undertaken to investigate the oocyte morphology, its fertilizing capacity and granulosa cell functions in ewes (obese, normal, metabolic stressed and emaciated). Ewes (Ovis aries) of approximately 3 years of age (Bellary breed) from a local village were screened, chosen and categorized into a) normal b) obese but not metabolically stressed, c) Emaciated but not metabolically stressed d) Metabolically stressed based on body condition scoring and blood markers. Oocytes and granulosa cells were collected from ovaries of the ewes of all categories after slaughter and were classified into good (oocytes with more than three layers of cumulus cells and homogenous ooplasm), fair (oocytes one or two layers of cumulus cells and homogenous ooplasm) and poor (denuded oocytes or with dark ooplasm). The good and fair quality oocytes were in vitro matured and cultured with fresh semen present and the fertilization, cleavage and blastocyst development were observed. The granulosa cells were cultured for evaluation of metabolic activity by use of the MTT assay, and cell viability, cell number as well as estrogen and progesterone production were assessed. It was observed that the good and fair quality oocytes had greater metabolic activity when collected from normal and obese ewes compared with those from emaciated and metabolically stressed ewes. No significant difference was observed in oocyte quality and maturation amongst the oocytes collected from normal and obese ewes. The cleavage and blastocyst production rates were different for the various body condition classifications and when ranked were: normal>obese>metabolically stressed>emaciated. Lesser metabolic activity was observed in granulosa cells obtained from ovaries of emaciated ewes. However, no changes were observed in viability and cell number of granulosa cells obtained from ewes with the different body condition categories. Estrogen and progesterone production from cultured granulosa cells were

  2. Granulosa cell tumors of the ovary : The clinical value of serum inhibin A and B levels in a large single center cohort

    NARCIS (Netherlands)

    Mom, C. H.; Engelen, M. J. A.; Willemse, P. H. B.; Gietema, J. A.; ten Hoor, K. A.; de Vries, E. G. E.; van der Zee, A. G. J.

    Objectives. In patients with a granulosa cell tumor of the ovary, the value of serum inhibin A and B concentrations for the assessment of disease status was investigated. Methods. In 30 consecutive patients with a stage I-III granulosa cell tumor, inhibin A and B concentrations were measured in pre-

  3. Cell cycle evaluation of granulosa cells in the {gamma}-irradiated mouse ovarian follicles

    Energy Technology Data Exchange (ETDEWEB)

    KIm, Jin Kyu; Lee, Chang Joo; Lee, Young Keun [Korea Atomic Energy Research Institute, Taejon (Korea, Republic of); Song, Kang Won; Yoon, Yong Dal [Hanyang Univ., Seoul (Korea, Republic of)

    1999-03-01

    This study was carried out to evaluate the biochemical and morphological effects of ionizing radiation on mouse ovarian follicles. Immature mice (ICR, 3 week-old) were irradiated with a dose of LD{sub 80(30)} at KAERI. The ovaries were collected after 6 hours, 12 hours, 1 day, and 2 days post irradiation. With the morphological basis of the histological staining with hematoxylin-eosin, immunohistochemical preparation using in situ 3'-end labeling was evaluated. Flow cytometric evaluation of DNA extracted from the whole ovary was performed. The percentage of A{sub 0} (subpopulation of cells with degraded DNA and with lower DNA fluorescence than G{sub 0}/G{sub 1} cells), apoptotic, cells in the cell cycle was significantly higher in the irradiated group than in the control group. The number of in situ 3'-end labeled follicles increased at 6 hours post irradiation. All the analyses represented that the ionizing radiation-induced follicular atresia was taken place via an apoptotic degeneration. Such a degeneration underwent very fast and acutely. Therefore, it is concluded that the radiation-induced follicular degeneration is, like the spontaneous atresia, mediated by an acute apoptosis of follicular granulosa cells. Flow cytometric evaluation of cell cycles can make the role for quantifying the atretic follicles and understanding the mechanism of the radiation-induced cell death.

  4. 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.

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

    Directory of Open Access Journals (Sweden)

    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.

  6. THE EFFECT OF GREEN TEA EXTRACT - EPIGALLOCATECHIN GALLATE (EGCG ON PORCINE OVARIAN GRANULOSA CELL

    Directory of Open Access Journals (Sweden)

    Attila Kádasi

    2014-02-01

    Full Text Available The aim of our study was to elucidate the potential effect of green tea substance on basic ovarian functions. For this purpose, we examined the action of green tea bioactive molecule, epigallocatechin gallate (given at doses 0, 1, 10, 100 μg/mL, on cultured porcine ovarian granulosa cell functions - proliferation, apoptosis and steroidogenesis. Accumulation of PCNA (marker of proliferation, BAX (marker of apoptosis and the release of steroid hormones (progesterone and testosterone were analysed by immunocytochemistry and RIA respectively. It was observed that epigallocatechin gallate addition decreased the percentage of proliferative (PCNA-positive cells at all used doses (1, 10 and 100 μg/mL. The percentage of apoptotic (BAX-positive cells was increased at the highest used dose (100 μg/mL, but not a lower doses. Epigallocatechin gallate stimulated progesterone release (at 10 μg/mL but not at 1 and 100 μg/mL and diminished testosterone release (at 1 μg/mL but not at 10 and 100 μg/mL by porcine granulosa cells. Our results suggest a direct effect of epigallocatechin gallate on proliferation, apoptosis and steroidogenesis in porcine ovaries. Taken together, these data suggest that green tea molecule epigallocatechin gallate can negatively affect reproductive (ovarian functions – suppress ovarian cell proliferation, promote their apoptosis and alter release of steroid hormones.

  7. 3,4-Dihydroxybenzaldehyde Derived from Prunus mume Seed Inhibits Oxidative Stress and Enhances Estradiol Secretion in Human Ovarian Granulosa Tumor Cells

    International Nuclear Information System (INIS)

    Kono, Ryohei; Nomura, Sachiko; Okuno, Yoshiharu; Nakamura, Misa; Maeno, Akihiro; Kagiya, Tomoko; Tokuda, Akihiko; Inada, Ken-ichi; Matsuno, Akira; Utsunomiya, Tomoko; Utsunomiya, Hirotoshi

    2014-01-01

    Granulosa cells form ovarian follicles and play important roles in the growth and maturation of oocytes. The protection of granulosa cells from cellular injury caused by oxidative stress is an effective therapy for female infertility. We here investigated an effective bioactive compound derived from Prunus mume seed extract that protects granulosa cells from hydrogen peroxide (H 2 O 2 )-induced apoptosis. We detected the bioactive compound, 3,4-dihydroxybenzaldehyde (3,4-DHBA), via bioactivity-guided isolation and found that it inhibited the H 2 O 2 -induced apoptosis of granulosa cells. We also showed that 3,4-DHBA promoted estradiol secretion in granulosa cells and enhanced the mRNA expression levels of steroidogenic factor 1, a promoter of key steroidogenic enzymes. These results suggest that P. mume seed extract may have clinical potential for the prevention and treatment of female infertility

  8. 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 (pascorbic acid might be involved in the regulation of selected biochemical and physiological processes in ovarian granulosa cells.

  9. Association between expression of cumulus expansion markers and real-time proliferation of porcine follicular granulosa cells in a primary cell culture model.

    Science.gov (United States)

    Ciesiółka, S; Budna, J; Bryja, A; Kranc, W; Chachuła, A; Dyszkiewicz-Konwińska, M; Piotrowska, H; Bukowska, D; Antosik, P; Bruska, M; Brüssow, K P; Nowicki, M; Zabel, M; Kempisty, B

    2016-01-01

    Folliculogenesis is a compound process that involves both ovarian follicle growth and oocyte development, which is tightly attached to the follicular wall. During this process, cells that form the follicle structure undergo substantial morphological and molecular modifications that finally lead to differentiation and specialization of ovarian follicular cells. The differentiation of ovarian cells encompasses formation of follicle, which is composed of theca (TCs), mural granulosa (GCs), and cumulus cells (CCs). It was previously hypothesized that GCs and CCs represent undifferentiated and highly specialized follicular cells, respectively, which may have similar primordial cell origins. In this study, we investigated the expression pattern of cumulus expansion markers such as COX2, HAS2, PTX3, and TSG6 in porcine GCs during short-term, in vitro culture. We hypothesized that these genes may display an important function in GCs in relation to cellular real-time proliferation. The expression pattern of COX2, HAS2, PTX3, and TSG6 was evaluated after using RT-qPCR in relation to confocal microscopy observations of protein expression and distribution during real-time proliferation of porcine follicular GCs. The COX2 and HAS2 mRNAs were highly expressed after 120 h of in vitro culture (IVC), whereas PTX3 and TSG6 mRNAs were increased during the first 24-48 h of IVC (P less than 0.001, P less than 0.01). Conversely, all of the encoded proteins were highly expressed after 144-168 h of IVC as compared to other culture periods (P less than 0.001, P less than 0.01). When analyzing the realtime proliferation of GCs in vitro, we observed a logarithmic increase of cell proliferation between 0 h and 120 h of IVC. However, after 120-168 h of IVC, the cells reached the lag phase of proliferation. Since it is well accepted that porcine GCs undergo luteinization shortly after 24-48 h of IVC, the expression pattern of investigated genes indicated that Cox2 and Has2 are independent from

  10. Effects of the mycotoxin deoxynivalenol on steroidogenesis and apoptosis in granulosa cells.

    Science.gov (United States)

    Guerrero-Netro, Hilda M; Chorfi, Younès; Price, Christopher A

    2015-06-01

    Mycotoxins can reduce fertility and development in livestock, notably in pigs and poultry, although the effect of most mycotoxins on reproductive function in cattle has not been established. One major mycotoxin, deoxynivalenol (DON), not only targets immune cells and activates the ribotoxic stress response (RSR) involving MAPK activation, but also inhibits oocyte maturation in pigs. In this study, we determined the effect of DON on bovine granulosa cell function using a serum-free culture system. Addition of DON inhibited estradiol and progesterone secretion, and reduced levels of mRNA encoding estrogenic (CYP19A1) but not progestogenic (CYP11A1 and STAR) proteins. Cell apoptosis was increased by DON, which also increased FASLG mRNA levels. The mechanism of action of DON was assessed by western blotting and PCR experiments. Addition of DON rapidly and transiently increased phosphorylation of MAPK3/1, and resulted in a more prolonged phosphorylation of MAPK14 (p38) and MAPK8 (JNK). Activation of these pathways by DON resulted in time- and dose-dependent increases in abundance of mRNA encoding the transcription factors FOS, FOSL1, EGR1, and EGR3. We conclude that DON is deleterious to granulosa cell function and acts through a RSR pathway. © 2015 Society for Reproduction and Fertility.

  11. Effect of GnRHa ovulation trigger dose on follicular fluid characteristics and granulosa cell gene expression profiles

    DEFF Research Database (Denmark)

    Vuong, Thi Ngoc Lan; Ho, M T; Ha, T Q

    2017-01-01

    in oocyte donors undergoing a single stimulation cycle at IVFMD, My Duc Hospital, Ho Chi Minh City, Vietnam, from August 2014 to March 2015. A total of 165 women aged 18-35 years with body mass index 1.25 ng/mL, and antral follicle count ≥6 were randomised to three...... granulosa cells were investigated in a subset of women from each group. RESULTS: Progesterone and oestradiol levels in FF did not differ significantly by trigger doses; findings were similar for 3βHSD, LHR and INHB-A gene expression in both cumulus and mural granulosa cells. CONCLUSIONS: In women co...

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

    Directory of Open Access Journals (Sweden)

    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.

  13. 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.

  14. In Vitro toxicological effects of Fumonisin B1 and Beauvericin on bovine granulosa cells

    Directory of Open Access Journals (Sweden)

    Marco Albonico

    2015-07-01

    Full Text Available Fumonisin B1 (FB1 and beauvericin (BEA are fusariotoxins found to co-exist in food and feed commodities. The aim of this study is to evaluate the individual and combined effects of FB1 and BEA on bovine granulosa cell proliferation and steroid production. Granulosa cells (GC from small bovine follicles (1-5 mm were cultured for 48 hours in 10% fetal bovine serum followed by 48 hours in a serum-free medium containing 500 ng/ml of testosterone (as an estradiol precursor, 30 ng/ml of FSH and 30 ng/ml of IGF-I with and without FB1 (3 µM and BEA (3 µM. At the end of the experiment, the numbers of GC were determined using a Coulter counter (Beckman Coulter, USA and concentrations of progesterone and estradiol in the culture medium were determined by radioimmunoassay. FB1 and BEA, both individually and in combination, showed an inhibitory effect (P 0.05 on estradiol and progesterone production, whereas BEA (3 µM, both alone and in combination with FB1 (3 µM, was found to decrease (P < 0.001 the production of both steroids drastically. In conclusion, this in vitro study indicates that FB1 and BEA, both individually and in combination, may affect GC proliferation to different extents and shows the drastic inhibitory effects of BEA on steroid production.

  15. Time- and Dose-Dependent Effects of 17 Beta-Estradiol on Short-Term, Real-Time Proliferation and Gene Expression in Porcine Granulosa Cells

    Directory of Open Access Journals (Sweden)

    Sylwia Ciesiółka

    2017-01-01

    Full Text Available The key mechanisms responsible for achievement of full reproductive and developmental capability in mammals are the differentiation and transformation of granulosa cells (GCs during folliculogenesis, oogenesis, and oocyte maturation. Although the role of 17 beta-estradiol (E2 in ovarian activity is widely known, its effect on proliferative capacity, gap junction connection (GJC formation, and GCs-luteal cells transformation requires further research. Therefore, the goal of this study was to assess the real-time proliferative activity of porcine GCs in vitro in relation to connexin (Cx, luteinizing hormone receptor (LHR, follicle stimulating hormone receptor (FSHR, and aromatase (CYP19A1 expression during short-term (168 h primary culture. The cultured GCs were exposed to acute (at 96 h of culture and/or prolonged (between 0 and 168 h of culture administration of 1.8 and 3.6 μM E2. The relative abundance of Cx36, Cx37, Cx40, Cx43, LHR, FSHR, and CYP19A1 mRNA was measured. We conclude that the proliferation capability of GCs in vitro is substantially associated with expression of Cxs, LHR, FSHR, and CYP19A1. Furthermore, the GC-luteal cell transformation in vitro may be significantly accompanied by the proliferative activity of GCs in pigs.

  16. Enhanced proliferation and progesterone production by porcine granulosa cells cultured with pseudorabies virus growth factor (PRGF).

    Science.gov (United States)

    Piekło, R; Gregoraszczuk, E L; Lesko, J; Golais, F; Stokłosowa, S

    1999-03-01

    The objective of this research was to study possible interactions of pseudorabies virus growth factor (PRGF) with ovarian tissue. Granulosa cells isolated from porcine ovaries were cultured as monolayers for 6 days in a control medium without PRGF and in medium supplemented with different doses of this agent. Increased population density and change towards more fibroblastic-like shape of cells cultured with 10(9) I.U PRGF was observed when compared with control culture. The cells divided significantly faster during 6 days of culture under the influence of 10(3), 10(4), 10(5), 10(6), 10(7), 10(8) and 10(9) I.U./ml of PRGF at a dose dependent manner. PRGF in a dose 10(9) I.U. added to cultured cells isolated from small and medium follicles did not influence progesterone secretion . An increase of progesterone secretion under the influence of PRGF in all investigated days of cultures was observed in cells isolated from large preovulatory follicles. The marked increase in progesterone content in PRGF treated culture in doses of 0.5x10(7), 0.5x10(8), 0.5x10(9) I.U. was observed during 4 and 6 days of culture. The rise of progesterone content was not connected with increased number of secretory cells, but with a stimulation of production per cell. PRGF exerted no visible effect on progesterone secretion by granulosa cells from small and medium follicles cultured for 6 days. The presented in vitro data provide evidence for a local action of PRGF in the follicle depending on the stage of follicular development and duration of exposure. Precise relevance of the interaction of PRGF with follicular development requires further study.

  17. 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.

  18. Vastatins have a distinct effect on sterol synthesis and progesterone secretion in human granulosa cells in vitro

    NARCIS (Netherlands)

    Vliet, A.K. van; Thiel, G.C.F. van; Naaktgeboren, N.; Cohen, L.H.

    1996-01-01

    Lovastatin and simvastatin are strong inhibitors of cholesterol synthesis in cultured human granulosa cells, as measured within 6 days after isolation, with IC50-values of respectively 27.0 and 18.2 nM obtained after 3.5 hours of incubation with the drugs. Pravastatin is a much weaker inhibitor of

  19. 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

  20. Evaluation of response to hormone therapy in patients with measurable adult granulosa cell tumors of the ovary

    NARCIS (Netherlands)

    van Meurs, Hannah S.; van der Velden, Jacobus; Buist, Marrije R.; van Driel, Willemien J.; Kenter, Gemma G.; van Lonkhuijzen, Luc R. C. W.

    2015-01-01

    The aim of this study was to retrospectively determine the objective response rate to hormone therapy (HT) for patients with a measurable adult granulosa cell tumor (GCT) of the ovary in a consecutive series of patients. All patients with an adult GCT who were treated with HT [steroidal progestins,

  1. Testosterone stimulates progesterone production and STAR, P450 cholesterol side-chain cleavage and LH receptor mRNAs expression in hen (Gallus domesticus) granulosa cells.

    Science.gov (United States)

    Rangel, P L; Rodríguez, A; Rojas, S; Sharp, P J; Gutierrez, C G

    2009-12-01

    The chicken ovary is organized into a hierarchy of yellow yolky follicles that ovulate on successive days. Active or passive immunization of laying hens against testosterone blocks ovulation without affecting follicle development. Testosterone may play a role in pre-ovulatory follicle maturation by stimulating granulosa progesterone production. We assessed whether this stimulus is dose-related and depends on the maturity of the donor follicle, and if it does so by stimulating granulosa cell STAR, P450 cholesterol side-chain cleavage (P450scc), and LH receptor (LHCGR) mRNAs expression. Progesterone production by granulosa cells from F1, F3, and F4 follicles, cultured for 3 h without testosterone was greater in cells collected 11-14 h than 1-4 h after ovulation. These differences in progesterone production were less pronounced after granulosa cells had been cultured for 24 h. Culture of granulosa cells for 3 or 24 h with testosterone (1-100 ng/ml) stimulated progesterone production in cells collected from F4, F3, or F1 follicles 1-4, or 11-14 h after ovulation. Testosterone (0-4000 ng/ml) alone or in combination with LH (0-100 ng/ml) increased progesterone production by F1 granulosa cells, collected 1-4 and 11-14 h after ovulation and cultured for 3 h. Finally, testosterone (10 or 100 ng/ml) increased STAR, P450scc, and LHCGR mRNAs, when added to 3 h cultures of F1 granulosa cells. In conclusion, testosterone stimulates granulosa cell progesterone production in hen pre-ovulatory hierarchical follicles irrespective of maturational state, acting alone or additively with LH. We propose that testosterone promotes granulosa cell maturation to facilitate the pre-ovulatory release of LH.

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

    Directory of Open Access Journals (Sweden)

    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.

  3. 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.

  4. The anti-Müllerian hormone (AMH) acts as a gatekeeper of ovarian steroidogenesis inhibiting the granulosa cell response to both FSH and LH.

    Science.gov (United States)

    Sacchi, Sandro; D'Ippolito, Giovanni; Sena, Paola; Marsella, Tiziana; Tagliasacchi, Daniela; Maggi, Elena; Argento, Cindy; Tirelli, Alessandra; Giulini, Simone; La Marca, Antonio

    2016-01-01

    Anti Müllerian Hormone (AMH) has a negative and inhibitory role in many functions of human granulosa-lutein cells (hGCs) including notoriously the reduction of the aromatase CYP19A1 expression induced by follicle-stimulating hormone (FSH). No data have been provided on the possible role of AMH in modulating the response to luteinizing hormone (LH) (alone or combined with FSH) as well as its effect on other enzymes involved in steroidogenesis including aromatase P450scc. The aim of this study was to investigate the role of AMH as regulator of the basal and stimulated steroids production by hGCs. Primary culture of hGCs were incubated with hormones AMH, LH, and FSH, alone or in combination. The CYP19A1 and P450scc messenger RNA (mRNA) expression, normalized by housekeeping ribosomal protein S7 (RpS7) gene, was evaluated by reverse transcriptase quantitative PCR (RT-qPCR). Each reaction was repeated in triplicate. Negative controls using corresponding amount of vehicle control for each hormone treatment were performed. AMH did not modulate the basal mRNA expression of both aromatase genes at any of the concentrations tested. Meanwhile, the strong mRNA induction of CYP19A1 and P450scc generated by a 24-h gonadotropin treatment (alone and combined) was suppressed by 20 ng/ml AMH added to culture medium. These findings contribute in clarifying the relationship between hormones regulating the early phase of steroidogenesis confirming that AMH is playing a suppressive role on CYP19A1 expression stimulated by gonadotropin in hGCs. Furthermore, a similar inhibitory effect for AMH was observed on P450scc gene expression when activated by gonadotropin treatment.

  5. Modulation of steroidogenesis by vitamin D3 in granulosa cells of the mouse model of polycystic ovarian syndrome.

    Science.gov (United States)

    Bakhshalizadeh, Shabnam; Amidi, Fardin; Alleyassin, Ashraf; Soleimani, Masoud; Shirazi, Reza; Shabani Nashtaei, Maryam

    2017-06-01

    Polycystic ovarian syndrome (PCOS) is the most common endocrine disorder of women of reproductive age characterized by polycystic ovarian morphology, anovulation or oligomenorrhea, and hyperandrogenism. It is shown that disruption in the steroidogenesis pathway caused by excess androgen in PCOS is a critical element of abnormal folliculogenesis and failure in dominant follicle selection. Vitamin D plays an important role in the regulation of ovulatory dysfunction and can influence genes involved in steroidogenesis in granulosa cells. In the present study, we investigated the effects of vitamin D3 on steroidogenic enzyme expression and activities in granulosa cell using a PCOS mouse model. In our study, the PCOS mouse model was developed by the injection of dehydroepiandrosterone (DHEA) for 20 days. The mRNA and protein expression levels of genes involved in steroidogenesis in granulosa cells were compared between polycystic and normal ovaries using real-time PCR and Western blotting assays. Granulosa cells of DHEA-induced PCOS mice were then cultured with and without vitamin D3 and mRNA and protein expression levels of steroidogenic enzymes and serum 17beta-estradiol and progesterone levels were investigated using qRT-PCR, western blot, and radioimmunoassay, respectively. Steroidogenic enzymes including Cyp11a1, StAR, Cyp19a1, and 3β-HSD were upregulated in granulosa cells of PCOS mice when compared to normal mice. Treatment with vitamin D3 decreased mRNA and protein expression levels of steroidogenic enzymes in cultured granulosa cells. Vitamin D3 also decreased aromatase and 3β-HSD activity that leads to decreased 17beta-estradiol and progesterone release. This study suggests that vitamin D3 could modulate the steroidogenesis pathway in granulosa cells of PCOS mice that may lead to improving follicular development and maturation. This is a step towards a possible conceivable treatment for PCOS. AMHR-II: anti-müllerian hormone receptor-II; 3β-HSD: 3

  6. Identification of potential biomarkers in donor cows for in vitro embryo production by granulosa cell transcriptomics

    DEFF Research Database (Denmark)

    Mazzoni, Gianluca; Salleh, Suraya M; Freude, Kristine

    2017-01-01

    The Ovum Pick Up-In vitro Production (OPU-IVP) of embryos is an advanced reproductive technology used in cattle production but the complex biological mechanisms behind IVP outcomes are not fully understood. In this study we sequenced RNA of granulosa cells collected from Holstein cows at oocyte......) involved in these mechanisms. We found a range of evidence that good IVP outcome is positively correlated with early follicular atresia. Furthermore we showed that high genetic index bulls can be used in breeding without reducing the IVP performances. These findings can contribute to the development......, TNFAIP6 and TXNDC11 were negatively associated while Mx1 and STC1 were positively associated with all IVP scores. Functional analysis highlighted a wide range of biological mechanisms including apoptosis, cell development and proliferation and four key upstream regulators (COX2, IL1, PRL, TRIM24...

  7. Effect of triptolide on progesterone production from cultured rat granulosa cells.

    Science.gov (United States)

    Zhang, J; Jiang, Z; Mu, X; Wen, J; Su, Y; Zhang, L

    2012-06-01

    Triptolide(CAS 38748-32-2), a major active component of Tripterygium wilfordii Hook F (TWHF), is known to have multiple pharmacological activities. However, studies have also shown that triptolide is highly disrupt to the reproductive system by disrupting normal steroid hormone signaling. In the present study, we investigated the effect of triptolide (5, 10, or 20 nM for 24 h) on progesterone production by rat granulosa cells. Triptolide inhibited both basal and human chorionic gonadotropin (HCG)- and 8-bromo-cAMP-stimulated progesterone production as revealed by RIA assay. Furthermore, the HCG-evoked increase in cellular cAMP content was also inhibited by triptolide, indicating that disruption of the cAMP/PKA signaling pathway may mediate the deleterious effects of triptolide on progesterone regulation. In addition, triptolide inhibited 25-OH-cholesterol-stimulated progesterone production, suggesting that activity of the P450 side chain cleavage (P450scc) enzyme was also be inhibited by triptolide. Western blot and quantitative real-time PCR (qRT-PCR) assays further revealed that triptolide decreased mRNA and protein expression of P450scc and the steroidogenic regulatory (StAR) protein in granulosa cells. In contrast, cell viability tests using 3-(4,5-dimethyl-thiazol-2-yl)-2,5- diphenyl-tetrazolium bromide (MTT) indicated that triptolide did not cause measurable cell death at doses that suppressed steroidogenesis. The reproductive toxicity of triptolide may be caused by disruption of cAMP/PKA-mediated expression of a number of progesterone synthesis enzymes or regulatory proteins, leading to reduced progesterone synthesis and reproductive dysfunction. © Georg Thieme Verlag KG Stuttgart · New York.

  8. Analysis of porcine granulosa cell death signaling pathways induced by vinclozolin.

    Science.gov (United States)

    Knet, Malgorzata; Wartalski, Kamil; Hoja-Lukowicz, Dorota; Tabarowski, Zbigniew; Slomczynska, Maria; Duda, Malgorzata

    2015-10-01

    Recent studies suggest that disturbing androgen-signaling pathways in porcine ovarian follicles may cause granulosa cell (GC) death. For this reason, we investigated which apoptotic pathway is initiated after GC exposure to an environmental antiandrogen, vinclozolin (Vnz), in vitro. Immunocytochemistry, Western blots, and fluorometric assays were used to quantify caspase-3 and -9 expression and activity. To elucidate the specific mechanism of Vnz action and toxicity, GCs were assessed for viability, cytotoxicity, and apoptotic activity using the ApoTox-Glo Triplex Assay. To further determine the mechanism of GC death induced by Vnz, we used the Apoptosis Antibody Array Kit. In response to Vnz stimulus, we found an increased level of caspase-3 protein expression (P ≤ 0.001) and an increase in caspase-3 proteolytic activity (P ≤ 0.001), confirming that Vnz is a potent proapoptotic factor. The strong immunoreaction of caspase-9 after Vnz treatment (P ≤ 0.001) suggests that intrinsic mitochondrial apoptosis pathway was activated during GC death. On the other hand, caspase-8, being a part of the extrinsic receptor pathway, was also activated (P ≤ 0.001). Therefore, it is possible that Vnz induces porcine granulosal apoptosis also through a parallel pathway. Activation of these two pathways was confirmed by the Apoptosis Antibody Array Kit. In conclusion, it is possible that the intrinsic signaling pathway may not act as an initial trigger for GC apoptosis but might contribute to the amplification and propagation of apoptotic cell death in the granulosa layer after treatment with this antiandrogen. Moreover, Vnz disturbs the physiological process of programmed cell death. Consequently, this could explain why atretic follicles are rapidly removed and suggests that normal function of the ovarian follicle may be destroyed. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. Juvenile granulosa cell ovarian tumor: a case report and review of literature.

    Science.gov (United States)

    Sivasankaran, Sujatha; Itam, Paul; Ayensu-Coker, Leslie; Sanchez, Judith; Egler, Rachel A; Anderson, Matthew L; Brandt, Mary L; Dietrich, Jennifer E

    2009-10-01

    Juvenile granulosa cell tumors (JGCT) are rare ovarian tumors that frequently present with precocious puberty. Presentation in infants less than a year of age is also rare. We describe a 10-month-old infant who presented with both premature thelarche and adrenarche due to JGCT. Laboratory evaluation revealed classic elevation of estradiol and inhibin B, and less classic elevation of total and free testosterone. Oophorectomy and staging resulted in a diagnosis of Stage IA JGCT. Survival rates are >95% among patients diagnosed under 10 years of age. Tumor recurrence is rare but can occur as late as 48 months. Therefore, tumor surveillance is warranted for patients with even a Stage IA JGCT and involves monitoring serial inhibin B levels along with intermittent imaging.

  10. Metastatic Granulosa Cell Tumor of the Testis: Clinical Presentation and Management

    Directory of Open Access Journals (Sweden)

    Anand Mohapatra

    2016-01-01

    Full Text Available Granulosa cell tumors (GCTs of the testis are rare sex cord-stromal tumors that are present in both juvenile and adult subtypes. While most adult GCTs are benign, those that present with distant metastases manifest a grave prognosis. Treatments for aggressive GCTs are not well established. Options that have been employed in previous cases include retroperitoneal lymph node dissection (RPLND, radiation, chemotherapy, or a combination thereof. We describe the case of a 57-year-old man who presented with a painless left testicular mass and painful gynecomastia. Serum tumor markers (alpha fetoprotein, human chorionic gonadotropin, and lactate dehydrogenase and computed tomography of the chest and abdomen were negative. The patient underwent left radical orchiectomy. Immunohistochemical staining was consistent with a testicular GCT. He underwent a left-template laparoscopic RPLND which revealed 2/19 positive lymph nodes. Final pathological stage was IIA. He remains free of disease 32 months after surgery.

  11. Aromatase inhibitors - a viable option for recurrent granulosa cell tumour of ovary: overview and case report

    International Nuclear Information System (INIS)

    Munem, A.A.; Bahrani, B.A.; Mehdi, I.

    2012-01-01

    Granulosa cell tumour of the ovary in adults is a rare tumour of low malignant potential affecting middle aged peri or post menopausal patients. These tumours are often diagnosed at an early stage, due to their hormonally active nature. They, however, have unique distinguishing histologic features and behaviour of frequent and late local or systemic relapses. The diagnosis can be challenging with unusual presentations. There is high association of endometrial carcinoma. Surgery is the mainstay of management in early low risk disease, while radiotherapy and systemic platinum based chemotherapy are employed in higher stage with poor prognostic indices. Survival is good in early stage disease. Recurrent, progressive, and treatment refractory disease is not infrequent and poses management challenge. Endocrine manipulation and hormone treatment are employed in few cases with equivocal results, as reported in literature. We present a case of recurrent and treatment refractory GCT in a postmenopausal patient, managed by aromatase inhibitor Anastrozole with reasonable efficacy. (author)

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

    Directory of Open Access Journals (Sweden)

    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

  13. IGF-1 attenuates LPS induced pro-inflammatory cytokines expression in buffalo (Bubalus bubalis) granulosa cells.

    Science.gov (United States)

    Onnureddy, K; Ravinder; Onteru, Suneel Kumar; Singh, Dheer

    2015-03-01

    Interaction between immune and endocrine system is a diverse process influencing cellular function and homeostasis in animals. Negative energy balance (NEB) during postpartum period in dairy animals usually suppresses these systems resulting in reproductive tract infection and infertility. These negative effects could be due to competition among endocrine and immune signaling pathways for common signaling molecules. The present work studied the effect of IGF-1 (50 ng/ml) on LPS (1 μg/ml) mediated pro-inflammatory cytokine expression (IL-1β, TNF-α, IL-6) and aromatase (CYP19A1) genes' expressions as well as proliferation of buffalo granulosa cells. The crosstalk between LPS and IGF-1 was also demonstrated through studying the activities of downstream signaling molecules (ERK1/2, Akt, NF-κB) by western blot and immunostaining. Gene expression analysis showed that IGF-1 significantly reduced the LPS induced expression of IL-1β, TNF-α and IL-6. LPS alone inhibited the CYP19A1 expression. However, co-treatment with IGF-1 reversed the inhibitory effect of LPS on CYP19A1 expression. LPS alone did not affect granulosa cell proliferation, but co-treatment with IGF-1, and IGF-1 alone enhanced the proliferation. Western blot results demonstrated that LPS caused the nuclear translocation of the NF-κB and increased the phosphorylation of ERK1/2 and Akt maximum at 15 min and 60 min, respectively. Nonetheless, co-treatment with IGF-1 delayed LPS induced phosphorylation of ERK1/2 (peak at 120 min), while promoting early Akt phosphorylation (peak at 5 min) with no effect on NF-κB translocation. Overall, IGF-1 delayed and reversed the effects of LPS, suggesting that high IGF-1 levels may combat infection during critical periods like NEB in postpartum dairy animals. Copyright © 2014 Elsevier Ltd. All rights reserved.

  14. 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, S. G.

    2011-01-01

    significantly with the expression of AMHRII, but did not correlate with any of the hormones in the follicular fluid. 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 follicular......Human small antral follicles (diameter 3-9 mm) were obtained from ovaries surgically removed for fertility preservation. From the individual aspirated follicles, granulosa cells and the corresponding follicular fluid were isolated in 64 follicles, of which 55 were available for mRNA analysis (24...... and to the follicular fluid concentrations of AMH, inhibin-B, progesterone and estradiol. AR mRNA expression in granulosa cells and the follicular fluid content of androgens both showed a highly significant positive association with the expression of FSHR mRNA in granulosa cells. AR mRNA expression also correlated...

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

    OpenAIRE

    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. ...

  16. Addition of granulosa cell mass to the culture medium of oocytes derived from early antral follicles increases oocyte growth, ATP content, and acetylation of H4K12.

    Science.gov (United States)

    Sugiyama, Miyako; Sumiya, Mei; Shirasuna, Koumei; Kuwayama, Takehito; Iwata, Hisataka

    2016-12-01

    The main aim of the present study was to examine the hypothesis that an increase in the number of granulosa cells surrounding developing bovine oocytes results in both high ATP levels and an increase in the acetylation level of H4K12 in oocytes grown in vitro. Oocyte-granulosa cell complexes (OGCs) were collected from early antral follicles (EAFs, 0.4-0.7 mm in diameter), and individually cultured on 96-well plates with or without additional granulosa cell mass that had been prepared from other OGCs. After 16 days of culture, we examined: (i) the rate of antrum formation of the OGCs; (ii) the diameter, maturation, and fertilization rate of the oocytes; and (iii) the ATP content and acetylation level of H4K12 in the oocytes grown in vitro. Granulosa cell mass added to the culture medium contributed to the development of OGCs with a higher rate of antrum formation and oocyte growth. Furthermore, the addition of granulosa cells increased the ATP content and acetylation level of H4K12 in oocytes grown in vitro compared with those developed without addition of granulosa cells. In addition, there was a positive correlation between the ATP content in oocytes grown in vitro and the number of granulosa cells in the corresponding OGCs. The results suggest that granulosa cells play a role not only in the development of OGCs and the growth of oocytes, but also in the determination of ATP content and the acetylation of H4K12 in the oocytes developed in vitro.

  17. Mucinous cystadenoma of the ovary with stromal luteinization and hilar cell hyperplasia during pregnancy.

    Science.gov (United States)

    Pascal, R R; Grecco, L A

    1988-02-01

    A 32-year-old woman was delivered of a healthy, full-term infant by cesarean section, at which time a large ovarian cyst was removed. The cyst proved to be a mucinous cystadenoma with prominent luteinization of the stroma subtending the epithelium and with numerous foci of hyperplastic Leydig cells in the cyst wall and ovarian hilum. These hormonally induced changes must be recognized in order to avoid mistaking them for invasive epithelial components.

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

    International Nuclear Information System (INIS)

    Ramasharma, K.; Li, C.H.

    1987-01-01

    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 α-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

  19. 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.

  20. Lysosomes are involved in induction of steroidogenic acute regulatory protein (StAR) gene expression and progesterone synthesis through low-density lipoprotein in cultured bovine granulosa cells.

    Science.gov (United States)

    Zhang, Jin-You; Wu, Yi; Zhao, Shuan; Liu, Zhen-Xing; Zeng, Shen-Ming; Zhang, Gui-Xue

    2015-09-15

    Progesterone is an important steroid hormone in the regulation of the bovine estrous cycle. The steroidogenic acute regulatory protein (StAR) is an indispensable component for transporting cholesterol to the inner mitochondrial membrane, which is one of the rate-limiting steps for progesterone synthesis. Low-density lipoprotein (LDL) supplies cholesterol precursors for progesterone formation, and the lysosomal degradation pathway of LDL is essential for progesterone biosynthesis in granulosa cells after ovulation. However, it is currently unknown how LDL and lysosomes coordinate the expression of the StAR gene and progesterone production in bovine granulosa cells. Here, we investigated the role of lysosomes in LDL-treated bovine granulosa cells. Our results reported that LDL induced expression of StAR messenger RNA and protein as well as expression of cholesterol side-chain cleavage cytochrome P-450 (CYP11A1) messenger RNA and progesterone production in cultured bovine granulosa cells. The number of lysosomes in the granulosa cells was also significantly increased by LDL; whereas the lysosomal inhibitor, chloroquine, strikingly abolished these LDL-induced effects. Our results indicate that LDL promotes StAR expression, synthesis of progesterone, and formation of lysosomes in bovine granulosa cells, and lysosomes participate in the process by releasing free cholesterol from hydrolyzed LDL. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. Transcriptomic Analysis and Meta-Analysis of Human Granulosa and Cumulus Cells.

    Directory of Open Access Journals (Sweden)

    Tanja Burnik Papler

    Full Text Available Specific gene expression in oocytes and its surrounding cumulus (CC and granulosa (GC cells is needed for successful folliculogenesis and oocyte maturation. The aim of the present study was to compare genome-wide gene expression and biological functions of human GC and CC. Individual GC and CC were derived from 37 women undergoing IVF procedures. Gene expression analysis was performed using microarrays, followed by a meta-analysis. Results were validated using quantitative real-time PCR. There were 6029 differentially expressed genes (q < 10-4; of which 650 genes had a log2 FC ≥ 2. After the meta-analysis there were 3156 genes differentially expressed. Among these there were genes that have previously not been reported in human somatic follicular cells, like prokineticin 2 (PROK2, higher expressed in GC, and pregnancy up-regulated nonubiquitous CaM kinase (PNCK, higher expressed in CC. Pathways like inflammatory response and angiogenesis were enriched in GC, whereas in CC, cell differentiation and multicellular organismal development were among enriched pathways. In conclusion, transcriptomes of GC and CC as well as biological functions, are distinctive for each cell subpopulation. By describing novel genes like PROK2 and PNCK, expressed in GC and CC, we upgraded the existing data on human follicular biology.

  2. Effect of adiponectin on bovine granulosa cell steroidogenesis, oocyte maturation and embryo development

    Directory of Open Access Journals (Sweden)

    Coyral-Castel Stéphanie

    2010-03-01

    Full Text Available Abstract Background Adiponectin is an adipokine, mainly produced by adipose tissue. It regulates several reproductive processes. The protein expression of the adiponectin system (adiponectin, its receptors, AdipoR1 and AdipoR2 and the APPL1 adaptor in bovine ovary and its role on ovarian cells and embryo, remain however to be determined. Methods Here, we identified the adiponectin system in bovine ovarian cells and embryo using RT-PCR, immunoblotting and immunohistochemistry. Furthermore, we investigated in vitro the effects of recombinant human adiponectin (10 micro g/mL on proliferation of granulosa cells (GC measured by [3H] thymidine incorporation, progesterone and estradiol secretions measured by radioimmunoassay in the culture medium of GC, nuclear oocyte maturation and early embryo development. Results We show that the mRNAs and proteins for the adiponectin system are present in bovine ovary (small and large follicles and corpus luteum and embryo. Adiponectin, AdipoR1 and AdipoR2 were more precisely localized in oocyte, GC and theca cells. Adiponectin increased IGF-1 10(-8 M-induced GC proliferation (P Conclusions In bovine species, adiponectin decreased insulin-induced steroidogenesis and increased IGF-1-induced proliferation of cultured GC through a potential involvement of ERK1/2 MAPK pathway, whereas it did not modify oocyte maturation and embryo development in vitro.

  3. 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.)

  4. 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.

  5. CHEMERIN (RARRES2) decreases in vitro granulosa cell steroidogenesis and blocks oocyte meiotic progression in bovine species.

    Science.gov (United States)

    Reverchon, Maxime; Bertoldo, Michael J; Ramé, Christelle; Froment, Pascal; Dupont, Joëlle

    2014-05-01

    CHEMERIN, or RARRES2, is a new adipokine that is involved in the regulation of adipogenesis, energy metabolism, and inflammation. Recent data suggest that it also plays a role in reproductive function in rats and humans. Here we studied the expression of CHEMERIN and its three receptors (CMKLR1, GPR1, and CCRL2) in the bovine ovary and investigated the in vitro effects of this hormone on granulosa cell steroidogenesis and oocyte maturation. By RT-PCR, immunoblotting, and immunohistochemistry, we found CHEMERIN, CMKLR1, GPR1, and CCRL2 in various ovarian cells, including granulosa and theca cells, corpus luteum, and oocytes. In cultured bovine granulosa cells, INSULIN, IGF1, and two insulin sensitizers-metformin and rosiglitazone-increased rarres2 mRNA expression whereas they decreased cmklr1, gpr1, and cclr2 mRNA expression. Furthermore, TNF alpha and ADIPONECTIN significantly increased rarres2 and cmklr1 expression, respectively. In cultured bovine granulosa cells, human recombinant CHEMERIN (hRec, 200 ng/ml) reduced production of both progesterone and estradiol, cholesterol content, STAR abundance, CYP19A1 and HMGCR proteins, and the phosphorylation levels of MAPK3/MAPK1 in the presence or absence of FSH (10(-8) M) and IGF1 (10(-8) M). All of these effects were abolished by using an anti-CMKLR1 antibody. In bovine cumulus-oocyte complexes, the addition of hRec (200 ng/ml) in the maturation medium arrested most oocytes at the germinal vesicle stage, and this was associated with a decrease in MAPK3/1 phosphorylation in both oocytes and cumulus cells. Thus, in cultured bovine granulosa cells, hRec decreases steroidogenesis, cholesterol synthesis, and MAPK3/1 phosphorylation, probably through CMKLR1. Moreover, in cumulus-oocyte complexes, it blocked meiotic progression at the germinal vesicle stage and inhibited MAPK3/1 phosphorylation in both the oocytes and cumulus cells during in vitro maturation. © 2014 by the Society for the Study of Reproduction, Inc.

  6. 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

    2016-06-08

    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.

  7. Epithelialization and stromalization of porcine follicular granulosa cells during real-time proliferation - a primary cell culture approach.

    Science.gov (United States)

    Ciesiółka, S; Bryja, A; Budna, J; Kranc, W; Chachuła, A; Bukowska, D; Piotrowska, H; Porowski, L; Antosik, P; Bruska, M; Brüssow, K P; Nowicki, M; Zabel, M; Kempisty, B

    2016-01-01

    The process of oocyte growth and development takes place during long stages of folliculogenesis and oogenesis. This is accompanied by biochemical and morphological changes, occurring from the preantral to antral stages during ovarian follicle differentiation. It is well known that the process of follicle growth is associated with morphological modifications of theca (TCs) and granulosa cells (GCs). However, the relationship between proliferation and/or differentiation of porcine GCs during long-term in vitro culture requires further investigation. Moreover, the expression of cytokeratins and vimentin in porcine GCs, in relation to real-time cell proliferation, has yet to be explored. Utilizing confocal microscopy, we analyzed cytokeratin 18 (CK18), cytokeratin 8 + 18 + 19 (panCK), and vimentin (Vim) expression, as well as their protein distribution, within GCs isolated from slaughtered ovarian follicles. The cells were cultured for 168 h with protein expression and cell proliferation index analyzed at 24-h intervals. We found the highest expression of CK18, panCK, and Vim occurred at 120 h of in vitro culture (IVC) as compared with other experimental time intervals. All of the investigated proteins displayed cytoplasmic distribution. Analysis of real-time cell proliferation revealed an increased cell index after the first 24 h of IVC. Additionally, during each period between 24-168 h of IVC, a significant difference in the proliferation profile, expressed as the cell index, was also observed. We concluded that higher expression of vimentin at 120 h of in vitro proliferation might explain the culmination of the stromalization process associated with growth and domination of stromal cells in GC culture. Cytokeratin expression within GC cytoplasm confirms the presence of epithelial cells as well as epithelial-related GC development during IVC. Moreover, expression of both cytokeratins and vimentin during short-term culture suggests that the process of GC proliferation

  8. Effect of intraovarian factors on porcine follicular cells: cumulus expansion, granulosa and cumulus cell progesterone production

    Czech Academy of Sciences Publication Activity Database

    Ježová, M.; Scsuková, S.; Nagyová, Eva; Vranová, J.; Procházka, Radek; Kolena, J.

    2001-01-01

    Roč. 65, - (2001), s. 115-126 ISSN 0378-4320 R&D Projects: GA ČR GA524/98/0231; GA AV ČR KSK5052113 Keywords : pig-ovary * cumulus expansion * luteinization stimulator Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.196, year: 2001

  9. Transcriptome profiling of sheep granulosa cells and oocytes during early follicular development obtained by Laser Capture Microdissection

    Directory of Open Access Journals (Sweden)

    Bonnet Agnes

    2011-08-01

    Full Text Available Abstract Background Successful achievement of early folliculogenesis is crucial for female reproductive function. The process is finely regulated by cell-cell interactions and by the coordinated expression of genes in both the oocyte and in granulosa cells. Despite many studies, little is known about the cell-specific gene expression driving early folliculogenesis. The very small size of these follicles and the mixture of types of follicles within the developing ovary make the experimental study of isolated follicular components very difficult. The recently developed laser capture microdissection (LCM technique coupled with microarray experiments is a promising way to address the molecular profile of pure cell populations. However, one main challenge was to preserve the RNA quality during the isolation of single cells or groups of cells and also to obtain sufficient amounts of RNA. Using a new LCM method, we describe here the separate expression profiles of oocytes and follicular cells during the first stages of sheep folliculogenesis. Results We developed a new tissue fixation protocol ensuring efficient single cell capture and RNA integrity during the microdissection procedure. Enrichment in specific cell types was controlled by qRT-PCR analysis of known genes: six oocyte-specific genes (SOHLH2, MAEL, MATER, VASA, GDF9, BMP15 and three granulosa cell-specific genes (KL, GATA4, AMH. A global gene expression profile for each follicular compartment during early developmental stages was identified here for the first time, using a bovine Affymetrix chip. Most notably, the granulosa cell dataset is unique to date. The comparison of oocyte vs. follicular cell transcriptomes revealed 1050 transcripts specific to the granulosa cell and 759 specific to the oocyte. Functional analyses allowed the characterization of the three main cellular events involved in early folliculogenesis and confirmed the relevance and potential of LCM-derived RNA. Conclusions

  10. The effect of yucca on proliferation, apoptosis, and steroidogenesis of porcine ovarian granulosa cells

    Directory of Open Access Journals (Sweden)

    Aneta Štochmaľová

    2014-02-01

    Full Text Available Yucca shidigera is a medicinal plant native to Mexico. Is a plant widely used in folk medicine to treat a variety of ailmentary disorders, but its action on reproductive processes and possible mechanisms of such action remains unknown. Yucca schidigera extract contains a number of steroidal saponins that, because of their biological activity, have attracted attention from the food industry for many years. Yucca extract is used as a natural feed additive with positive effect to microflora, digestion, metabolism and to improve animal muscle growth. Its extract has been used as a foodstuff and folk medicine to treat a wide variety of diseases for many years. Nevertheless, it remaines unknown, whether consumption of yucca can affect reproductive system. The aim of this study was to examine the effects of yucca on basic ovarian cell functions - proliferation, apoptosis and steroidogenesis. Porcine ovarian granulosa cells were cultured with and without yucca extract (added at doses 0; 1; 10 and 100 μg.mL-1 of medium. Markers of proliferation (% of PCNA-positive cells and apoptosis (% cells containing bax were analysed by immunocytochemistry. Release of steroid hormones (progesterone and testosterone was measured by EIA. It was observed, that addition of yucca inhibited proliferation (expression of PCNA, increased apoptosis (expression of bax, stimulated progesterone and inhibited testosterone release. The ability of yucca to reduce ovarian cell proliferation, to promote ovarian cell apoptosis and affect steroidogenesis demonstrates the direct influence of yucca on female gonads. Furthermore, our observations suggest the multiple sites of action (proliferation, apoptosis, steroidogenesis of yucca on porcine ovarian cell functions. It is not to be excluded, that consumption of yucca can suppress female reproductive functions.

  11. GATA4 and GATA6 Knockdown During Luteinization Inhibits Progesterone Production and Gonadotropin Responsiveness in the Corpus Luteum of Female Mice.

    Science.gov (United States)

    Convissar, Scott M; Bennett, Jill; Baumgarten, Sarah C; Lydon, John P; DeMayo, Francesco J; Stocco, Carlos

    2015-12-01

    The surge of luteinizing hormone triggers the genomic reprogramming, cell differentiation, and tissue remodeling of the ovulated follicle, leading to the formation of the corpus luteum. During this process, called luteinization, follicular granulosa cells begin expressing a new set of genes that allow the resulting luteal cells to survive in a vastly different hormonal environment and to produce the extremely high amounts of progesterone (P4) needed to sustain pregnancy. To better understand the molecular mechanisms involved in the regulation of luteal P4 production in vivo, the transcription factors GATA4 and GATA6 were knocked down in the corpus luteum by crossing mice carrying Gata4 and Gata6 floxed genes with mice carrying Cre recombinase fused to the progesterone receptor. This receptor is expressed exclusively in granulosa cells after the luteinizing hormone surge, leading to recombination of floxed genes during follicle luteinization. The findings demonstrated that GATA4 and GATA6 are essential for female fertility, whereas targeting either factor alone causes subfertility. When compared to control mice, serum P4 levels and luteal expression of key steroidogenic genes were significantly lower in conditional knockdown mice. The results also showed that GATA4 and GATA6 are required for the expression of the receptors for prolactin and luteinizing hormone, the main luteotropic hormones in mice. The findings demonstrate that GATA4 and GATA6 are crucial regulators of luteal steroidogenesis and are required for the normal response of luteal cells to luteotropins. © 2015 by the Society for the Study of Reproduction, Inc.

  12. PGRMC1 participates in late events of bovine granulosa cells mitosis and oocyte meiosis.

    Science.gov (United States)

    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.

  13. Congenital juvenile granulosa cell tumor of the testis: Case report and literature review

    Directory of Open Access Journals (Sweden)

    Carolina Talini

    2016-07-01

    Full Text Available Juvenile granulosa cell tumor (JGCT is a very rarely diagnosed benign tumor, accounting for 1.2% of all prepubertal testicular tumors. A full-term healthy neonate was diagnosed with a painless left scrotal mass. During evaluation it was identified to have about two times the volume of the contralateral testis, presenting a firm consistency, not as hard as the consistency of a prenatal testicular torsion. Doppler ultrasound detected a multicystic left testicular mass, with normal blood flow, but failed in detecting normal-appearing testis. Human chorionic gonadotropin (β-HCG and serum alpha-fetoprotein (AFP were normal. Inguinal approach was performed, section of the lesion was sent to frozen biopsy and excluded yolk sac tumor, and however the impossibility of detecting normal testis tissue indicated orchiectomy with high ligation of the spermatic cord. Histological evaluation demonstrated gray testicular parenchyma with multicystic aspect fulfilled with yellow fluid. The usual clinical presentation of JGCT is a painless scrotal mass, radiological imaging demonstrates a multicystic tumor. Tumoral markers levels are normal and the standard treatment is the inguinal orchiectomy.

  14. Three-dimensional culture of buffalo granulosa cells in hanging drop mimics the preovulatory follicle stage.

    Science.gov (United States)

    Yadav, Monica; Agrawal, Himanshu; Pandey, Mamta; Singh, Dheer; Onteru, Suneel K

    2018-03-01

    Granulosa cell (GC) culture models mimicking the intrafollicular environment are limited. Such models have a great potential in reproductive toxicity studies. The buffalo, a monovulatory species like humans, could be a better model than polyovulatory rodents. Therefore, we targeted the development and characterization of three-dimensional (3D) culture systems for buffalo GCs. The GCs from small ovarian follicles (SF) maintained the CYP19 gene expression for 144 hr in a 2D culture system. Hence, GCs from SF were cultured directly in 3D using hanging drop and Poly-([2-hydroxyethyl methacrylate]) (polyHEMA) methods in the DMEM media containing 1 ng/ml FSH and 10 ng/ml IGF-1 for 144 hr. The expression profile of nine GC-specific transcripts; CYP19, TNFAIP6, AMH, PTI, NR4A1, FSHR, RUNX, LHR, and COX2/PTGS2; revealed that 3D-spheroids developed in hanging drop method maintained the GC phenotype of preovulatory follicles. Therefore, hanging drop method is a best method for culturing GCs to mimic the intrafollicular environment. © 2017 Wiley Periodicals, Inc.

  15. Role of adjuvant radiotherapy in granulosa cell tumors of the ovary.

    Science.gov (United States)

    Hauspy, Jan; Beiner, Mario E; Harley, Ian; Rosen, Barry; Murphy, Joan; Chapman, William; Le, Lisa W; Fyles, Anthony; Levin, Wilfred

    2011-03-01

    To review the role of adjuvant radiotherapy (RT) in the outcome and recurrence patterns of granulosa cell tumors (GCTs) of the ovary. The records of all patients with GCTs referred to the Princess Margaret Hospital University Health Network between 1961 and 2006 were retrospectively reviewed. The patient, tumor, and treatment factors were assessed by univariate and multivariate analyses using disease-free survival (DFS) as the endpoint. A total of 103 patients with histologically confirmed GCTs were included in the present study. The mean duration of follow-up was 100 months (range, 1-399). Of the 103 patients, 31 received adjuvant RT. A total of 39 patients developed tumor recurrence. The tumor size, incidence of intraoperative rupture, and presence of concurrent endometrial cancer were not significant risk factors for DFS. The median DFS was 251 months for patients who underwent adjuvant RT compared with 112 months for patients who did not (p=.02). On multivariate analysis, adjuvant RT remained a significant prognostic factor for DFS (p=.004). Of the 103 patients, 12 had died and 44 were lost to follow-up. Ovarian GCTs can be indolent, with patients achieving long-term survival. In our series, adjuvant RT resulted in a significantly longer DFS. Ideally, randomized trials with long-term follow-up are needed to define the role of adjuvant RT for ovarian GCTs. Crown Copyright © 2011. Published by Elsevier Inc. All rights reserved.

  16. 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. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  17. Simultaneous effects of endocrine disruptor bisphenol A and flavonoid fisetin on progesterone production by granulosa cells.

    Science.gov (United States)

    Bujnakova Mlynarcikova, Alzbeta; Scsukova, Sona

    2018-04-01

    In the present study, we aimed to examine effects of different concentrations of the endocrine disruptor Bisphenol A (BPA; 1 nM, 1 μM, 100 μM) and the flavonoid fisetin (1, 10, 25, 50 μM), individually and in combinations, on steroidogenic function of porcine ovarian granulosa cells (GCs) represented by progesterone production. We confirmed that BPA inhibited progesterone production by GCs at the highest concentration. Fisetin reduced gonadotropin-stimulated progesterone synthesis dose-dependently, and in this manner, fisetin impaired progesterone production when added to BPA-treated GCs. The mechanisms of the inhibitory effects of the combinations included a significant down-regulation of the key steroidogenesis-related genes (STAR, CYP11A1, HSD3B). Our findings suggest for the first time that fisetin might interfere with ovarian steroidogenesis, and might not have beneficial but rather aggravating effects in terms of modulating progesterone synthesis altered by high concentrations of BPA. Copyright © 2018 Elsevier B.V. All rights reserved.

  18. Expression of endothelin-1 gene and protein in human granulosa cells

    Energy Technology Data Exchange (ETDEWEB)

    Magini, A.; Granchi, S.; Susini, T. [Univ. of Florence (Italy)] [and others

    1996-04-01

    Previous studies in animal models indicated an autocrine/paracrine action of endothelin-1 (ET-1) in the ovary. We now report evidence on the presence of ET-1 in human ovary during reproductive life. Immunohistochemical and in situ hybridization studies demonstrated a positive signal into cytoplasm of granulosa cells (GC) of follicles at different growth stages. The concentration of ET-1-like immunoreactivity (ET-1-Li) was also measured by a specific RIA in human follicular fluid (FF). FF samples were obtained from women in an in vitro fertilization program undergoing gonadotropin stimulation (group A; n = 24) or no treatment (group B; n = 7). The mean ({+-}SD) ET-1-LI FF level in group A (4.85 {+-} 2.06 pg/mL) was significantly higher than that in group B (1.29 {+-} 0.43 pg/mL; P < 0.01), whereas the corresponding mean plasma levels were not significantly different and were not correlated to respective FF values. Our results indicate for the first time the presence of ET-1 and its messenger ribonucleic acid in the GC of the human ovary. The higher ET-1-LI levels found in the FF from women undergoing gonadotropin treatment suggest a modulation by gonadotropins and/or ovarian steroids of ET-1 production by GC. 19 refs., 4 figs., 1 tab.

  19. 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.

  20. Prohibitin (PHB) acts as a potent survival factor against ceramide induced apoptosis in rat granulosa cells.

    Science.gov (United States)

    Chowdhury, Indrajit; Branch, Alicia; Olatinwo, Moshood; Thomas, Kelwyn; Matthews, Roland; Thompson, Winston E

    2011-08-29

    Ceramide is a key factor in inducing germ cell apoptosis by translocating from cumulus cells into the adjacent oocyte and lipid rafts through gap junctions. Therefore studies designed to elucidate the mechanistic pathways in ceramide induced granulosa cell (GC) apoptosis and follicular atresia may potentially lead to the development of novel lipid-based therapeutic strategies that will prevent infertility and premature menopause associated with chemo and/or radiation therapy in female cancer patients. Our previous studies have shown that Prohibitin (PHB) is intimately involved in GCs differentiation, atresia, and luteolysis. In the present study, we have examined the functional effects of loss-/gain-of-function of PHB using adenoviral technology in delaying apoptosis induced by the physiological ligand ceramide in rat GCs. Under these experimental conditions, exogenous ceramide C-8 (50 μM) augmented the expression of mitochondrial PHB and subsequently cause the physical destruction of GC by the release of mitochondrial cytochrome c and activation of caspase-3. In further studies, silencing of PHB expression by adenoviral small interfering RNA (shRNA) sensitized GCs to ceramide C8-induce apoptosis. In contrast, adenovirus (Ad) directed overexpression of PHB in GCs resulted in increased PHB content in mitochondria and delayed the onset of ceramide induced apoptosis in the infected GCs. Taken together, these results provide novel evidences that a critical level of PHB expression within the mitochondria plays a key intra-molecular role in GC fate by mediating the inhibition of apoptosis and may therefore, contribute significantly to ceramide induced follicular atresia. Copyright © 2011 Elsevier Inc. All rights reserved.

  1. Tributyltin or triphenyltin inhibits aromatase activity in the human granulosa-like tumor cell line KGN.

    Science.gov (United States)

    Saitoh, M; Yanase, T; Morinaga, H; Tanabe, M; Mu, Y M; Nishi, Y; Nomura, M; Okabe, T; Goto, K; Takayanagi, R; Nawata, H

    2001-11-23

    The superimposition of male sex organs (penis and vas deferens) in a female gastropod, called imposex, is widely attributed to the exposure to tributyltin (TBT) compounds, used world-wide in antifouling paints for ships. It has been hypothesized that the TBT-induced imposex is mediated by an increasing androgen level relative to the estrogen level, namely a decreased conversion of androgens to estrogens (i.e., aromatization). In the present study, we tested this hypothesis by examining the effects of TBT or triphenyltin (TPT) on the aromatase activity in a cultured human granulosa-like tumor cell line, KGN, which was recently established by our group. Treatment with more than 1000 ng/ml TBT compounds was very toxic to the cells and caused immediate cell death within 24 h, while 200 ng/ml was found to cause apoptosis of the cells. Treatment of the KGN cells for more than 48 h with 20 ng/ml TBT or TPT, which is a concentration level reported to cause imposex in marine species, did not affect cell proliferation but significantly suppressed the aromatase activity determined by a [(3)H]H(2)O release assay. Treatment with 20 ng/ml TBT compounds for 7 days also resulted in a reduction of the E2 production from Delta 4-androstenedione stimulated by db-cAMP. The changes in the aromatase activity by TBT compounds were associated with comparable changes in P450arom mRNA assessed by RT-PCR. The luciferase activity of the P450arom promoter II (1 kb) decreased after the addition of 20 ng/ml TBT compounds in transfected KGN cells either in a basic state or in states stimulated by db-cAMP. The Ad4BP-dependent increase in the luciferase activity of P450arom promoter II was also downregulated by such treatments. These results indicate that TBT compounds inhibited the aromatase activity and also decreased the P450arom mRNA level at the transcriptional level in KGN cells. The direct inhibitory effect of TBT compounds on the aromatase activity may therefore partly explain the induction

  2. TGF-β1 downregulates StAR expression and decreases progesterone production through Smad3 and ERK1/2 signaling pathways in human granulosa cells.

    Science.gov (United States)

    Fang, Lanlan; Chang, Hsun-Ming; Cheng, Jung-Chien; Leung, Peter C K; Sun, Ying-Pu

    2014-11-01

    Regulation of progesterone production in granulosa cells is important for normal reproductive functions. Steroidogenic acute regulatory protein (StAR) is recognized as the key regulatory protein involved in the rate-limiting step of steroidogenesis. TGF-β1 protein is detected in human follicular fluid, and TGF-β1 and its receptors are expressed in human granulosa cells. However, the functional role of TGF-β1 in the regulation of StAR expression and progesterone production in human granulosa cells remains unknown. Our objective was to investigate the effects of TGF-β1 on StAR expression and progesterone production in human granulosa cells. SVOG cells are human granulosa cells that were obtained from women undergoing in vitro fertilization and immortalized with SV40 large T antigen. SVOG cells were used to investigate the effects of TGF-β1 on StAR expression and progesterone production at an academic research center. Levels of mRNA and protein were examined by RT-qPCR and western blotting, respectively. The accumulation levels of progesterone were measured by enzyme-linked immunosorbent assay (ELISA). TGF-β1 treatment downregulated StAR expression and decreased progesterone production. The suppressive effects of TGF-β1 on StAR expression and progesterone production were abolished by the inhibition of TGF-β type I receptor. In addition, treatment with TGF-β1 activated the Smad2/3 and ERK1/2 signaling pathways. The inhibition of the Smad3 and ERK1/2 signaling pathways attenuated the TGF-β1-induced downregulation of StAR expression and progesterone production. TGF-β1 downregulated StAR expression and decreased progesterone production by activating the Smad3 and ERK1/2 signaling pathways in human granulosa cells.

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

    Directory of Open Access Journals (Sweden)

    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.

  4. Targeted genome-wide DNA methylation profiling of ovarian granulosa cells from women with PCOS

    Directory of Open Access Journals (Sweden)

    Pooja Sagvekar

    2017-10-01

    Full Text Available Polycystic ovary syndrome (PCOS is a complex endocrinopathy of obscure pathophysiologic origins, globally affecting 6-15% women of childbearing age. Emerging evidence on repercussions of environmental insults and changing lifestyles on fecundity and reproductive health have necessitated the study of tissue-specific epigenetic alterations in PCOS development. In semblance to follicular and oocyte defects observed in PCOS ovaries, targeted bisulfite sequencing was performed to generate the methylome signatures of ovarian granulosa cells (GCs obtained from age-BMI matched women with PCOS (n=3 and healthy, regularly menstruating controls (n=3 using next generation sequencing approach. Paired end sequencing of samples was carried out on Illumina HiSeq 2500 ® platform and data were analyzed using the Bismark tool. Methylation levels of a few selected genes relevant to ovarian function were further validated in GCs obtained from 10 controls and 10 women with PCOS by pyrosequencing.  Relative transcript levels of these genes were assessed by q-RT PCR using Taqman assays. In the methylome analysis, a total of 6486 CpG sites representing 3840 genes associated with pathways such as Wnt signaling, G-protein receptor signaling, angiogenesis, chemokine and cytokine mediated inflammation and integrin signaling showed differential methylation in PCOS. Of these, a total of 2977 CpG sites representing 2063 genes were identified as hypomethylated while 3509 CpG sites in 1777 genes were found to be hypermethylated. Additionally, differential methylation was also noted in several non-coding RNAs regulating vital ovarian functions and which are reported to be dysregulated in PCOS. This data provides compelling evidence in support of epigenetic alterations as etiopathogenic factors associated with ovarian dysfunction in PCOS.

  5. Prohibitin regulates the FSH signaling pathway in rat granulosa cell differentiation.

    Science.gov (United States)

    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. © 2016 The authors.

  6. miR-26b promotes granulosa cell apoptosis by targeting ATM during follicular atresia in porcine ovary.

    Directory of Open Access Journals (Sweden)

    Fei Lin

    Full Text Available More than 99% of ovarian follicles undergo atresia in mammals, but the mechanism of follicular atresia remains to be elucidated. In this study, we explored microRNA (miRNA regulation of follicular atresia in porcine ovary. A miRNA expression profile was constructed for healthy, early atretic, and progressively atretic follicles, and the differentially expressed miRNAs were selected and analyzed. We found that miR-26b, which was upregulated during follicular atresia, increased the number of DNA breaks and promoted granulosa cell apoptosis by targeting the ataxia telangiectasia mutated gene directly in vitro.

  7. The effect of metformin treatment in vivo on acute and long-term energy metabolism and progesterone production in vitro by granulosa cells from women with polycystic ovary syndrome.

    Science.gov (United States)

    Maruthini, D; Harris, S E; Barth, J H; Balen, A H; Campbell, B K; Picton, H M

    2014-10-10

    . Granulosa cells from women with PCOS pathology revealed reduced pyruvate production and preferential lactate production in addition to their reduced glucose uptake during cultures (P progesterone synthesis compared with controls (P progesterone biosynthetic capacity of the GCs from women with PCOS. The positive, acute metabolic effect and the negative long-term steroidogenic effect on GCs following metformin exposure in vivo may have important implications for follicular development and luteinized GC function when the drug is used in clinical practice. No competing interests. This work was supported by the UK Medical Research Council Grant Reference number G0800250. © The Author 2014. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology.

  8. Different molecular organization of two carotenoids, lutein and zeaxanthin, in human colon epithelial cells and colon adenocarcinoma cells

    Science.gov (United States)

    Grudzinski, Wojciech; Piet, Mateusz; Luchowski, Rafal; Reszczynska, Emilia; Welc, Renata; Paduch, Roman; Gruszecki, Wieslaw I.

    2018-01-01

    Two cell lines, human normal colon epithelial cells (CCD 841 CoTr) and human colon adenocarcinoma cells (HT-29) were cultured in the presence of exogenous carotenoids, either zeaxanthin or lutein. Both carotenoids demonstrated cytotoxicity with respect to cancer cells but not to normal cells. Cells from both the cell lines were analyzed with application of fluorescence lifetime imaging microscopy and Raman scattering microscopy. Both imaging techniques show effective incorporation of carotenoid molecules into growing cells. Comparison of the Raman scattering and fluorescence lifetime characteristics reveals different molecular organization of carotenoids in the carcinoma and normal cells. The main difference consists in a carotenoid aggregation level which is substantially lower in the carcinoma cells as compared to the normal cells. Different molecular organization of carotenoids was interpreted in terms of a different metabolism of normal and carcinoma cells and has been concluded to provide a possibility of cancer diagnosis based on spectroscopic analyses.

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

    Directory of Open Access Journals (Sweden)

    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

  10. Competence Classification of Cumulus and Granulosa Cell Transcriptome in Embryos Matched by Morphology and Female Age.

    Directory of Open Access Journals (Sweden)

    Rehannah Borup

    Full Text Available By focussing on differences in the mural granulosa cell (MGC and cumulus cell (CC transcriptomes from follicles resulting in competent (live birth and non-competent (no pregnancy oocytes the study aims on defining a competence classifier expression profile in the two cellular compartments.A case-control study.University based facilities for clinical services and research.MGC and CC samples from 60 women undergoing IVF treatment following the long GnRH-agonist protocol were collected. Samples from 16 oocytes where live birth was achieved and 16 age- and embryo morphology matched incompetent oocytes were included in the study.MGC and CC were isolated immediately after oocyte retrieval. From the 16 competent and non-competent follicles, mRNA was extracted and expression profile generated on the Human Gene 1.0 ST Affymetrix array. Live birth prediction analysis using machine learning algorithms (support vector machines with performance estimation by leave-one-out cross validation and independent validation on an external data set.We defined a signature of 30 genes expressed in CC predictive of live birth. This live birth prediction model had an accuracy of 81%, a sensitivity of 0.83, a specificity of 0.80, a positive predictive value of 0.77, and a negative predictive value of 0.86. Receiver operating characteristic analysis found an area under the curve of 0.86, significantly greater than random chance. When applied on 3 external data sets with the end-point outcome measure of blastocyst formation, the signature resulted in 62%, 75% and 88% accuracy, respectively. The genes in the classifier are primarily connected to apoptosis and involvement in formation of extracellular matrix. We were not able to define a robust MGC classifier signature that could classify live birth with accuracy above random chance level.We have developed a cumulus cell classifier, which showed a promising performance on external data. This suggests that the gene signature at

  11. 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

    2016-04-26

    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.

  12. Transcriptomal profiling of bovine ovarian granulosa and theca interna cells in primary culture in comparison with their in vivo counterparts.

    Directory of Open Access Journals (Sweden)

    Nicholas Hatzirodos

    Full Text Available In vitro culture of ovarian granulosa cells and theca cells has been very important for our understanding of their function and regulation. One of the most eagerly sought attributes of cell culture is the use of chemically-defined conditions. However, even under such in vitro conditions cell behaviour could differ from the in vivo situation because of differences in oxygen tension, nutrients, adhesion matrix and other factors. To examine this further we compared the transcriptomes of both granulosa cells and cells from the theca interna that were cultured in what are arguably the best in vitro conditions for maintaining the 'follicular' phenotypes of both tissue types, as displayed by their respective freshly-isolated counterparts. The array data analysed are from recently published data and use the same sizes of bovine follicles (small antral 3-6 mm and the same Affymetrix arrays. We conducted analysis using Partek, Ingenuity Pathway Analysis and GOEAST. Principal Component Analysis (PCA and hierarchical clustering clearly separated the in vivo from the in vitro groups for both cells types and transcriptomes were more homogeneous upon culture. In both cell cultures behaviours associated with cell adhesion, migration and interaction with matrix or substrate were more abundant. However, the pathways involved generally differed between the two cell types. With the thecal cultures a gene expression signature of an immune response was more abundant, probably by leukocytes amongst the cells cultured from the theca interna. These results indicate differences between in vivo and in vitro that should be considered when interpreting in vitro data.

  13. Identification of differential gene expression in in vitro FSH treated pig granulosa cells using suppression subtractive hybridization

    Directory of Open Access Journals (Sweden)

    Tosser-Klopp G

    2006-07-01

    Full Text Available Abstract FSH, which binds to specific receptors on granulosa cells in mammals, plays a key role in folliculogenesis. Its biological activity involves stimulation of intercellular communication and upregulation of steroidogenesis, but the entire spectrum of the genes regulated by FSH has yet to be fully characterized. In order to find new regulated transcripts, however rare, we have used a Suppression Subtractive Hybridization approach (SSH on pig granulosa cells in primary culture treated or not with FSH. Two SSH libraries were generated and 76 clones were sequenced after selection by differential screening. Sixty four different sequences were identified, including 3 novel sequences. Experiments demonstrated the presence of 25 regulated transcripts. A gene ontology analysis of these 25 genes revealed (1 catalytic; (2 transport; (3 signal transducer; (4 binding; (5 anti-oxidant and (6 structural activities. These findings may deepen our understanding of FSH's effects. Particularly, they suggest that FSH is involved in the modulation of peroxidase activity and remodelling of chromatin.

  14. Effects of Lutein on Hyperosmoticity-Induced Upregulation of IL-6 in Cultured Corneal Epithelial Cells and Its Relevant Signal Pathways

    Directory of Open Access Journals (Sweden)

    Shih-Chun Chao

    2016-01-01

    Full Text Available Dry eye is a common disorder characterized by deficiency of tear. Hyperosmoticity of tear stimulates inflammation and damage of ocular surface tissues and plays an essential role in the pathogenesis of dry eye. Cultured human corneal epithelial (CE cells were used for the study of effects of lutein and hyperosmoticity on the secretion of IL-6 by CE cells. Cell viability of CE cells was not affected by lutein at 1–10 μM as determined by MTT assay. Hyperosmoticity significantly elevated the secretion of IL-6 by CE cells as measured by ELISA analysis. The constitutive secretion of IL-6 was not affected by lutein. Lutein significantly and dose-dependently inhibited hyperosmoticity-induced secretion of IL-6. Phosphorylated- (p- p38 MAPK, p-JNK levels in cell lysates and NF-κB levels in cell nuclear extracts were increased by being exposed to hyperosmotic medium. JNK, p38, and NF-κB inhibitors decreased hyperosmoticity-induced secretion of IL-6. Lutein significantly inhibited hyperosmoticity-induced elevation of NF-κB, p38, and p-JNK levels. We demonstrated that lutein inhibited hyperosmoticity-induced secretion of IL-6 in CE cells through the deactivation of p38, JNK, and NF-κB pathways. Lutein may be a promising agent to be explored for the treatment of dry eye.

  15. Progesterone Receptor Membrane Component 1 (PGRMC1 in cell division: its role in bovine granulosa cells mitosis

    Directory of Open Access Journals (Sweden)

    Laura Terzaghi

    2015-07-01

    Full Text Available The present studies were aimed to assess Progesterone Receptor Membrane Component-1 (PGRMC1 role in regulating bovine granulosa cells (bGC mitosis. First, we performed immunofluorescence studies on in vitro cultured bGC collected from antral follicles, which showed that PGRMC1 localizes to the spindle apparatus in mitotic cells. Then, to evaluate PGRMC1 effect on cell proliferation we silenced its expression with RNA interference technique (RNAi. Quantitative RT-PCR and immunoblotting confirmed down-regulation of PGRMC1 expression, when compared to CTRL-RNAi treated bGC (p<0.05. After 72h of culture, PGRMC1 silencing determined a lower growth rate (p<0.05 and a higher percentage of cells arrested at G2/M phase as assessed by flowcytometry (p<0.05. Accordingly, live imaging studies revealed more aberrant mitosis and a delayed M-phase in PGRMC1-RNAi treated cells compared to CTRL-RNAi group (p<0.05. These data confirmed that PGRMC1 is directly involved in bGC mitosis and ongoing preliminary studies are aimed to elucidate its putative mechanisms of action. Since PGRMC1 is a membrane protein, we hypothesize its possible involvement in vesicular trafficking and endocytosis, which is in turn an important process to assure proper cell division. To assess this hypothesis, we have preliminarily conducted immunofluorescence and in situ proximity ligation assay experiments that showed PGRMC1 co-localization and direct interaction with clathrin. This is important since clathrin is an essential protein for both endosomes formation, and cell division acting directly on the spindle apparatus. Thus our studies set the stage for analysis aimed to further characterize PGRMC1’s mechanism of action in mitotic cell.

  16. 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

    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

  17. 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...

  18. Development and internal validation of a prognostic model to predict recurrence free survival in patients with adult granulosa cell tumors of the ovary

    NARCIS (Netherlands)

    van Meurs, Hannah S.; Schuit, Ewoud; Horlings, Hugo M.; van der Velden, Jacobus; van Driel, Willemien J.; Mol, Ben Willem J.; Kenter, Gemma G.; Buist, Marrije R.

    2014-01-01

    Models to predict the probability of recurrence free survival exist for various types of malignancies, but a model for recurrence free survival in individuals with an adult granulosa cell tumor (GCT) of the ovary is lacking. We aimed to develop and internally validate such a prognostic model. We

  19. Increased expression of pentraxin 3 after in vivo and in vitro stimulation with gonadotropins in porcine oocyte-cumulus complexes and granulosa cells

    Czech Academy of Sciences Publication Activity Database

    Nagyová, Eva; Kalous, Jaroslav; Němcová, Lucie

    2016-01-01

    Roč. 56, č. 1 (2016), s. 29-35 ISSN 0739-7240 R&D Projects: GA ČR GA305/05/0960 Institutional support: RVO:67985904 Keywords : oocyte-cumulus complex * granulosa cells * pentraxin 3 Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.644, year: 2016

  20. IL-1α Up-Regulates IL-6 Expression in Bovine Granulosa Cells via MAPKs and NF-κB Signaling Pathways

    Directory of Open Access Journals (Sweden)

    Meng Yang

    2017-01-01

    Full Text Available Background/Aims: IL-6 is one of the main cytokines in regulating ovarian follicular development and ovulation. However, the factors that regulate IL-6 expression in follicles are still unclear. The aim of this study was to elucidate the mechanisms underlying the effect of IL-1α on IL-6 expression in granulosa cells. Methods: IL-6 expression after IL-1α with/without inhibitors treatment was analyzed by RT-qPCR and ELISA. The phosphorylation of proteins induced by IL-1α was analyzed by western blot. The intracellular cAMP level was assayed by immunoassay kit. Results: IL-1α has a dose-dependent effect on IL-6 expression in granulosa cells. This promoting effect can be significantly attenuated by Erk, c-Jun, p38 and IκB proteins inhibitors, respectively. Moreover, the phosphorylation levels of Erk, c-Jun, p38 and IκBα proteins were significantly increased after IL-1α treatment. In addition, we also found that IL-1α not only reversed the cAMP attenuated IL-6 expression, but also increased IL-1α mRNA expression in granulosa cells. Conclusion: The regulation of IL-1α on IL-6 expression is mediated by activation of MAPKs and NF-κB signaling pathways. Moreover,IL-1α may regulate the ovulation-related genes expression in granulosa cells by an autocrine and/or paracrine manner.

  1. High fat diet triggers cell cycle arrest and excessive apoptosis of granulosa cells during the follicular development

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Yanqing; Zhang, Zhenghong; Liao, Xinghui; Wang, Zhengchao, E-mail: zcwang@fjnu.edu.cn

    2015-10-23

    The regulatory mechanism of granulosa cells (GCs) proliferation during the follicular development is complicated and multifactorial, which is essential for the oocyte growth and normal ovarian functions. To investigate the role of high fat diet (HFD) on the proliferation of GCs, 4-week old female mice were fed with HFD or normal control diet (NC) for 15 weeks or 20 weeks and then detected the expression level of some regulatory molecules of cell cycle and apoptosis. The abnormal ovarian morphology was observed at 20 weeks. Further mechanistic studies indicated that HFD induced-obesity caused elevated apoptotic levels in GCs of the ovaries in a time-dependent manner. Moreover, cell cycle progress was also impacted after HFD fed. The cell cycle inhibitors, p27{sup Kip1} and p21{sup Cip1}, were significantly induced in the ovaries from the mice in HFD group when compared with that in the ovaries from the mice in NC group. Subsequently, the expression levels of Cyclin D1, D3 and CDK4 were also significantly influenced in the ovaries from the mice fed with HFD in a time-dependent manner. The present results suggested that HFD induced-obesity may trigger cell cycle arrest and excessive apoptosis of GCs, causing the abnormal follicular development and ovarian function failure. - Highlights: • HFD induced-obesity leads to abnormal ovarian morphology. • HFD induced-obesity triggers excessive apoptosis in the ovary. • HFD induced-obesity up-regulates cell cycle inhibitors p21{sup Cip1} and p27{sup Kip1} in the ovary. • HFD induced-obesity causes cell cycle arrest in the ovary.

  2. MicroRNA-424/503 cluster members regulate bovine granulosa cell proliferation and cell cycle progression by targeting SMAD7 gene through activin signalling pathway.

    Science.gov (United States)

    Pande, Hari Om; Tesfaye, Dawit; Hoelker, Michael; Gebremedhn, Samuel; Held, Eva; Neuhoff, Christiane; Tholen, Ernst; Schellander, Karl; Wondim, Dessie Salilew

    2018-05-01

    The granulosa cells are indispensable for follicular development and its function is orchestrated by several genes, which in turn posttranscriptionally regulated by microRNAs (miRNA). In our previous study, the miRRNA-424/503 cluster was found to be highly abundant in bovine granulosa cells (bGCs) of preovulatory dominant follicle compared to subordinate counterpart at day 19 of the bovine estrous cycle. Other study also indicated the involvement of miR-424/503 cluster in tumour cell resistance to apoptosis suggesting this miRNA cluster may involve in cell survival. However, the role of miR-424/503 cluster in granulosa cell function remains elusive Therefore, this study aimed to investigate the role of miRNA-424/503 cluster in bGCs function using microRNA gain- and loss-of-function approaches. The role of miR-424/503 cluster members in granulosa cell function was investigated by overexpressing or inhibiting its activity in vitro cultured granulosa cells using miR-424/503 mimic or inhibitor, respectively. Luciferase reporter assay showed that SMAD7 and ACVR2A are the direct targets of the miRNA-424/503 cluster members. In line with this, overexpression of miRNA-424/503 cluster members using its mimic and inhibition of its activity by its inhibitor reduced and increased, respectively the expression of SMAD7 and ACVR2A. Furthermore, flow cytometric analysis indicated that overexpression of miRNA-424/503 cluster members enhanced bGCs proliferation by promoting G1- to S- phase cell cycle transition. Modulation of miRNA-424/503 cluster members tended to increase phosphorylation of SMAD2/3 in the Activin signalling pathway. Moreover, sequence specific knockdown of SMAD7, the target gene of miRNA-424/503 cluster members, using small interfering RNA also revealed similar phenotypic and molecular alterations observed when miRNA-424/503 cluster members were overexpressed. Similarly, to get more insight about the role of miRNA-424/503 cluster members in activin signalling

  3. Oleic acid induces specific alterations in the morphology, gene expression and steroid hormone production of cultured bovine granulosa cells.

    Science.gov (United States)

    Yenuganti, Vengala Rao; Viergutz, Torsten; Vanselow, Jens

    2016-06-01

    After parturition, one of the major problems related to nutritional management that is faced by the majority of dairy cows is negative energy balance (NEB). During NEB, excessive lipid mobilization takes place and hence the levels of free fatty acids, among them oleic acid, increase in the blood, but also in the follicular fluid. This accumulation can be associated with serious metabolic and reproductive disorders. In the present study, we analyzed the effects of physiological concentrations of oleic acid on cell morphology, apoptosis, necrosis, proliferation and steroid production, and on the abundance of selected transcripts in cultured bovine granulosa cells. Increasing oleic acid concentrations induced intracellular lipid droplet accumulation, thus resulting in a foam cell-like morphology, but had no effects on apoptosis, necrosis or proliferation. Oleic acid also significantly reduced the transcript abundance of the gonadotropin hormone receptors, FSHR and LHCGR, steroidogenic genes STAR, CYP11A1, HSD3B1 and CYP19A1, the cell cycle regulator CCND2, but not of the proliferation marker PCNA. In addition, treatment increased the transcript levels of the fatty acid transporters CD36 and SLC27A1, and decreased the production of 17-beta-estradiol and progesterone. From these data it can be concluded that oleic acid specifically affects morphological and physiological features and gene expression levels thus altering the functionality of granulosa cells. Suggestively, these effects might be partly due to the reduced expression of FSHR and thus the reduced responsiveness to FSH stimulation. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  4. C/EBPβ Promotes STAT3 Expression and Affects Cell Apoptosis and Proliferation in Porcine Ovarian Granulosa Cells.

    Science.gov (United States)

    Yuan, Xiaolong; Zhou, Xiaofeng; He, Yingting; Zhong, Yuyi; Zhang, Ailing; Zhang, Zhe; Zhang, Hao; Li, Jiaqi

    2018-06-13

    Previous studies suggest that signal transducer and activator of transcription 3 (STAT3) and CCAAT/enhancer binding protein beta (C/EBPβ) play an essential role in ovarian granulosa cells (GCs) for mammalian follicular development. Several C/EBPβ putative binding sites were previously predicted on the STAT3 promoter in mammals. However, the molecular regulation of C/EBPβ on STAT3 and their effects on cell proliferation and apoptosis remain virtually unexplored in GCs. Using porcine GCs as a model, the 5′-deletion, luciferase report assay, mutation, chromatin immunoprecipitation, Annexin-V/PI staining and EdU assays were applied to investigate the molecular mechanism for C/EBPβ regulating the expression of STAT3 and their effects on the cell proliferation and apoptosis ability. We found that over and interfering with the expression of C/EBPβ significantly increased and decreased the messenger RNA (mRNA) and protein levels of STAT3 , respectively. The dual luciferase reporter assay showed that C/EBPβ directly bound at −1397/−1387 of STAT3 to positively regulate the mRNA and protein expressions of STAT3 . Both C/EBPβ and STAT3 were observed to inhibit cell apoptosis and promote cell proliferation. Furthermore, C/EBPβ might enhance the antiapoptotic and pro-proliferative effects of STAT3 . These results would be of great insight in further exploring the molecular mechanism of C/EBPβ and STAT3 on the function of GCs and the development of ovarian follicles in mammals.

  5. 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.

  6. Toxicological effects of fumonisin B1 alone and in combination with other fusariotoxins on bovine granulosa cells.

    Science.gov (United States)

    Albonico, Marco; Schütz, Luis F; Caloni, Francesca; Cortinovis, Cristina; Spicer, Leon J

    2016-08-01

    There is now overwhelming evidence of global contamination of commodities with Fusarium mycotoxins. Fumonisin B1 (FB1) is a Fusarium mycotoxin frequently occurring in corn in combination with deoxynivalenol (DON), α-zearalenol (α-ZEA) and β-zearalenol (β-ZEA). The aim of this study was to determine if FB1, alone and combined with DON or α-ZEA or β-ZEA, can affect cell proliferation and steroid production of bovine granulosa cells (GC). A species-specific model with bovine granulosa cells (GC) was used to study the potential endocrine disruptor effects of FB1 alone and in co-exposure. In the presence of β-ZEA (30 ng/mL), FB1 at 30 ng/mL showed a stimulatory effect on GC numbers. Insulin-like growth factor-1 (IGF1)-stimulated cell proliferation was decreased after exposure to β-ZEA alone at 5.0 μg/mL and FB1 with α-ZEA and β-ZEA at the same concentration. Regarding steroid production, FB1 at 30 ng/mL and 100 ng/mL amplified the inhibitory effect of β-ZEA (30 ng/mL) on estradiol (E2) production, while FB1 alone increased (P < 0.05) IGF1-induced E2 production. α-ZEA alone decreased (P < 0.05) E2 production, whereas β-ZEA alone and in combination with FB1 decreased (P < 0.05) E2 production. These studies indicate for the first time that the Fusarium mycotoxin FB1 along with other mycotoxins can affect GC proliferation and steroid production, which ultimately could influence reproductive function in cattle. Copyright © 2016 Elsevier Ltd. All rights reserved.

  7. 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. © 2014 by the Society for the Study of Reproduction, Inc.

  8. Serum anti-Müllerian hormone concentrations before and after treatment of an ovarian granulosa cell tumour in a cat

    Directory of Open Access Journals (Sweden)

    Laura A Heaps

    2017-08-01

    Full Text Available Case summary A 15-year-old female cat was presented for investigation of progressive behavioural changes, polyuria, polydipsia and periuria. An ovarian granulosa cell tumour was identified and the cat underwent therapeutic ovariohysterectomy (OHE. The cat’s clinical signs resolved, but 6 months later it was diagnosed as having an anaplastic astrocytoma and was euthanased. Serum anti-Müllerian hormone (AMH concentration prior to OHE was increased vs a control group of entire and neutered female cats. Following OHE, serum AMH concentration decreased to <1% of the original value. Relevance and novel information Serum AMH measurement may represent a novel diagnostic and monitoring tool for functional ovarian neoplasms in cats.

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

    Directory of Open Access Journals (Sweden)

    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.

  10. Cultured bovine granulosa cells rapidly lose important features of their identity and functionality but partially recover under long-term culture conditions.

    Science.gov (United States)

    Yenuganti, Vengala Rao; Vanselow, Jens

    2017-05-01

    Cell culture models are essential for the detailed study of molecular processes. We analyze the dynamics of changes in a culture model of bovine granulosa cells. The cells were cultured for up to 8 days and analyzed for steroid production and gene expression. According to the expression of the marker genes CDH1, CDH2 and VIM, the cells maintained their mesenchymal character throughout the time of culture. In contrast, the levels of functionally important transcripts and of estradiol and progesterone production were rapidly down-regulated but showed a substantial up-regulation from day 4. FOXL2, a marker for granulosa cell identity, was also rapidly down-regulated after plating but completely recovered towards the end of culture. In contrast, expression of the Sertoli cell marker SOX9 and the lesion/inflammation marker PTGS2 increased during the first 2 days after plating but gradually decreased later on. We conclude that only long-term culture conditions (>4 days) allow the cells to recover from plating stress and to re-acquire characteristic granulosa cell features.

  11. Onset of lipoprotein-supported steroidogenesis in differentiating granulosa cells of rats: cellular events involved in mediating FSH-enhanced uptake of low-density lipoproteins

    International Nuclear Information System (INIS)

    Foster, J.D.

    1987-01-01

    Luteal cells use lipoproteins as the main source of cholesterol in steroidogenesis. However, little is known about the mechanisms underlying hormonal control of lipoprotein uptake. Thus, the authors tested the hypothesis that FSH and androgens regulate low density lipoprotein (LDL)-supported steroidogenesis in maturing granulosa cells by affecting receptor-mediated endocytosis of LDL at a cellular level. For this, immature ovarian granulosa cells were cultured with or without hormones, compactin (de novo synthesis inhibitor), or unlabeled or labeled ( 125 I or gold particles) LDL. Nonhormone-treated cultures produced little progestin; FSH and FSH/androstenedione stimulated steroid secretion. Progestin production by hormone-, but not nonhormone-, treated cultures was decreased by compactin, suggesting that de novo synthesis provided sterol for steroidogenesis. EM quantitation of cells exposed to gold-LDL at 37 0 C revealed that, compared to nonhormone-treated cells, FSH-treated cells (1) bound and internalized more gold-LDL, (2) had a smaller percentage of gold-LDL at their surfaces, (3) displayed a faster apparent rate of LDL internalization and delivery to lysosomes, and (4) contained more gold-labeled lysosomes. Data from biochemical studies in which 125 I-LDL was used supported the morphological findings. In conclusion, this study demonstrates that FSH has important effects at the cellular level on LDL uptake, which seem to underlie the striking increase in progestin production accompanying granulosa cell differentiation

  12. Constitutive luteinizing hormone receptor signaling causes sexual dysfunction and Leydig cell adenomas in male mice.

    Science.gov (United States)

    Hai, Lan; Hiremath, Deepak S; Paquet, Marilène; Narayan, Prema

    2017-05-01

    The luteinizing hormone receptor (LHCGR) is necessary for fertility, and genetic mutations cause defects in reproductive development and function. Activating mutations in LHCGR cause familial male-limited precocious puberty (FMPP). We have previously characterized a mouse model (KiLHRD582G) for FMPP that exhibits the same phenotype of precocious puberty, Leydig cell hyperplasia, and elevated testosterone as boys with the disorder. We observed that KiLHRD582G male mice became infertile by 6 months of age, although sperm count and motility were normal. In this study, we sought to determine the reason for the progressive infertility and the long-term consequences of constant LHCGR signaling. Mating with superovulated females showed that infertile KiLHRD582G mice had functional sperm and normal accessory gland function. Sexual behavior studies revealed that KiLHRD582G mice mounted females, but intromission was brief and ejaculation was not achieved. Histological analysis of the reproductive tract showed unique metaplastic changes resulting in pseudostratified columnar epithelial cells with cilia in the ampulla and chondrocytes in the penile body of the KiLHRD582G mice. The infertile KiLHRD582G exhibited enlarged sinusoids and a decrease in smooth muscle content in the corpora cavernosa of the penile body. However, collagen content was unchanged. Leydig cell adenomas and degenerating seminiferous tubules were seen in 1-year-old KiLHRD582G mice. We conclude that progressive infertility in KiLHRD582G mice is due to sexual dysfunction likely due to functional defects in the penis. © The Authors 2017. Published by Oxford University Press on behalf of Society for the Study of Reproduction. All rights reserved. For permissions, please journals.permissions@oup.com.

  13. Radical Scavenging Activity-Based and AP-1-Targeted Anti-Inflammatory Effects of Lutein in Macrophage-Like and Skin Keratinocytic Cells

    Directory of Open Access Journals (Sweden)

    Jueun Oh

    2013-01-01

    Full Text Available Lutein is a naturally occurring carotenoid with antioxidative, antitumorigenic, antiangiogenic, photoprotective, hepatoprotective, and neuroprotective properties. Although the anti-inflammatory effects of lutein have previously been described, the mechanism of its anti-inflammatory action has not been fully elucidated. Therefore, in the present study, we aimed to investigate the regulatory activity of lutein in the inflammatory responses of skin-derived keratinocytes or macrophages and to elucidate the mechanism of its inhibitory action. Lutein significantly reduced several skin inflammatory responses, including increased expression of interleukin-(IL- 6 from LPS-treated macrophages, upregulation of cyclooxygenase-(COX- 2 from interferon-γ/tumor necrosis-factor-(TNF- α-treated HaCaT cells, and the enhancement of matrix-metallopeptidase-(MMP- 9 level in UV-irradiated keratinocytes. By evaluating the intracellular signaling pathway and the nuclear transcription factor levels, we determined that lutein inhibited the activation of redox-sensitive AP-1 pathway by suppressing the activation of p38 and c-Jun-N-terminal kinase (JNK. Evaluation of the radical and ROS scavenging activities further revealed that lutein was able to act as a strong anti-oxidant. Taken together, our findings strongly suggest that lutein-mediated AP-1 suppression and anti-inflammatory activity are the result of its strong antioxidative and p38/JNK inhibitory activities. These findings can be applied for the preparation of anti-inflammatory and cosmetic remedies for inflammatory diseases of the skin.

  14. Insulin-like growth factors (IGFs) as autocrine/paracrine regulators of granulosa cell differentiation and growth: Studies with a neutralizing monoclonal antibody to IGF-I

    International Nuclear Information System (INIS)

    Mondschein, J.S.; Canning, S.F.; Miller, D.Q.; Hammond, J.M.

    1989-01-01

    Evidence that granulosa cells secrete and respond to insulin-like growth factors (IGFs) suggests, but does not prove, the importance of IGFs as intraovarian regulators. To further assess the role of these peptides in ovarian function, a neutralizing monoclonal antibody to IGF-I was employed to block the actions of IGFs in porcine follicular fluid and in granulosa cell-conditioned medium. In one series of experiments, granulosa cells from immature porcine follicles were cultured in medium containing porcine follicular fluid that had been charcoal-treated to remove steroids. As noted before, fluid from large follicles (LFF) stimulated progesterone production in a dose-dependent manner. The stimulatory effect of LFF (30% v/v) could be inhibited by greater than 50% by the anti-IGF monoclonal antibody. This inhibitory action was specific for the anti-IGF antibody and could be overcome by the addition of excess exogenous IGFs. In another series of experiments, granulosa cells were made dependent on endogenously produced IGFs by culturing them in a serum-free medium without exogenous growth factors. The effects of follicle-stimulating hormone (FSH), estradiol (E2), growth hormone (GH), and combinations thereof on progesterone production were inhibited by approximately 50% by the anti-IGF antibody. The effects of IGFs on indices of cell growth (judged by the criterion of being inhibited by the anti-IGF antibody) were less dramatic. A modest 18% increase in cell number was observed with FSH and E2 treatment in serum-free medium; this effect was virtually abolished by the antibody

  15. Insulin-like growth factors (IGFs) as autocrine/paracrine regulators of granulosa cell differentiation and growth: Studies with a neutralizing monoclonal antibody to IGF-I

    Energy Technology Data Exchange (ETDEWEB)

    Mondschein, J.S.; Canning, S.F.; Miller, D.Q.; Hammond, J.M. (Pennsylvania State Univ., Hershey (USA))

    1989-07-01

    Evidence that granulosa cells secrete and respond to insulin-like growth factors (IGFs) suggests, but does not prove, the importance of IGFs as intraovarian regulators. To further assess the role of these peptides in ovarian function, a neutralizing monoclonal antibody to IGF-I was employed to block the actions of IGFs in porcine follicular fluid and in granulosa cell-conditioned medium. In one series of experiments, granulosa cells from immature porcine follicles were cultured in medium containing porcine follicular fluid that had been charcoal-treated to remove steroids. As noted before, fluid from large follicles (LFF) stimulated progesterone production in a dose-dependent manner. The stimulatory effect of LFF (30% v/v) could be inhibited by greater than 50% by the anti-IGF monoclonal antibody. This inhibitory action was specific for the anti-IGF antibody and could be overcome by the addition of excess exogenous IGFs. In another series of experiments, granulosa cells were made dependent on endogenously produced IGFs by culturing them in a serum-free medium without exogenous growth factors. The effects of follicle-stimulating hormone (FSH), estradiol (E2), growth hormone (GH), and combinations thereof on progesterone production were inhibited by approximately 50% by the anti-IGF antibody. The effects of IGFs on indices of cell growth (judged by the criterion of being inhibited by the anti-IGF antibody) were less dramatic. A modest 18% increase in cell number was observed with FSH and E2 treatment in serum-free medium; this effect was virtually abolished by the antibody.

  16. Inhibitory effect of luteolin on estrogen biosynthesis in human ovarian granulosa cells by suppression of aromatase (CYP19).

    Science.gov (United States)

    Lu, Dan-feng; Yang, Li-juan; Wang, Fei; Zhang, Guo-lin

    2012-08-29

    Inhibition of aromatase, the key enzyme in estrogen biosynthesis, is an important strategy in the treatment of breast cancer. Several dietary flavonoids show aromatase inhibitory activity, but their tissue specificity and mechanism remain unclear. This study found that the dietary flavonoid luteolin potently inhibited estrogen biosynthesis in a dose- and time-dependent manner in KGN cells derived from human ovarian granulosa cells, the major source of estrogens in premenopausal women. Luteolin decreased aromatase mRNA and protein expression in KGN cells. Luteolin also promoted aromatase protein degradation and inhibited estrogen biosynthesis in aromatase-expressing HEK293A cells, but had no effect on recombinant expressed aromatase. Estrogen biosynthesis in KGN cells was inhibited with differing potencies by extracts of onion and bird chili and by four other dietary flavonoids: kaempferol, quercetin, myricetin, and isorhamnetin. The present study suggests that luteolin inhibits estrogen biosynthesis by decreasing aromatase expression and destabilizing aromatase protein, and it warrants further investigation as a potential treatment for estrogen-dependent cancers.

  17. No specific gene expression signature in human granulosa and cumulus cells for prediction of oocyte fertilisation and embryo implantation.

    Directory of Open Access Journals (Sweden)

    Tanja Burnik Papler

    Full Text Available In human IVF procedures objective and reliable biomarkers of oocyte and embryo quality are needed in order to increase the use of single embryo transfer (SET and thus prevent multiple pregnancies. During folliculogenesis there is an intense bi-directional communication between oocyte and follicular cells. For this reason gene expression profile of follicular cells could be an important indicator and biomarker of oocyte and embryo quality. The objective of this study was to identify gene expression signature(s in human granulosa (GC and cumulus (CC cells predictive of successful embryo implantation and oocyte fertilization. Forty-one patients were included in the study and individual GC and CC samples were collected; oocytes were cultivated separately, allowing a correlation with IVF outcome and elective SET was performed. Gene expression analysis was performed using microarrays, followed by a quantitative real-time PCR validation. After statistical analysis of microarray data, there were no significantly differentially expressed genes (FDR<0,05 between non-fertilized and fertilized oocytes and non-implanted and implanted embryos in either of the cell type. Furthermore, the results of quantitative real-time PCR were in consent with microarray data as there were no significant differences in gene expression of genes selected for validation. In conclusion, we did not find biomarkers for prediction of oocyte fertilization and embryo implantation in IVF procedures in the present study.

  18. 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 ...

  19. The effects of the environmental antiandrogen vinclozolin on the induction of granulosa cell apoptosis during follicular atresia in pigs.

    Science.gov (United States)

    Knet, Malgorzata; Tabarowski, Zbigniew; Slomczynska, Maria; Duda, Malgorzata

    2014-06-01

    The aim of this study was to investigate whether the androgens testosterone and dihydrotestosterone (DHT) and the antiandrogenic fungicide vinclozolin (Vnz) exert proapoptotic effects on porcine granulosa cells (GCs), and to examine the roles of these compounds in follicular atresia. Granulosa cells isolated from pig follicles were cultured for 24 hours, and then exposed to 0.1 μM testosterone, 0.1 μM DHT, 14 μM Vnz, or the equivalent concentrations of testosterone and Vnz or DHT and Vnz for a further 24 hours. Apoptosis and necrosis of the GCs were determined via Hoechst staining and flow cytometry analyses of annexin V-stained cells. Whole porcine follicles were also exposed to the same compounds and combinations of compounds for 24 hours. The sections were stained with hematoxylin and eosin for morphologic assessments, and a Terminal deoxynucleotidyl Transferase Biotyn-dUTP Nick-End Labeling (TUNEL) assay was performed to determine the number of apoptotic cells. The progesterone and estradiol concentrations secreted into the culture media by isolated GCs and follicles were also measured. Exposure to the androgens resulted in an increased number of apoptotic GCs both in vitro and in the organotypic model. Vinclozolin exposure increased and decreased the number of necrotic and apoptotic GCs, respectively. Furthermore, compared with control follicles, those exposed to testosterone, DHT, or Vnz displayed enhanced atresia, and coadministration of Vnz attenuated the promotive effect of these androgens on atresia. Estradiol secretion was stimulated by the combination of testosterone and Vnz, whereas exposure to Vnz alone reduced it. Progesterone production declined after the combined addition of androgens and the antiandrogen. In summary, Vnz caused massive necrosis of GCs in vitro and induced apoptosis of GCs in whole follicles. The androgens testosterone and DHT enhanced these effects. The results presented here suggest that selective destruction of porcine

  20. Expression pattern of G protein‑coupled estrogen receptor 1 (GPER) in human cumulus granulosa cells (CGCs) of patients with PCOS.

    Science.gov (United States)

    Zang, Lili; Zhang, Quan; Zhou, Yi; Zhao, Yan; Lu, Linlin; Jiang, Zhou; Peng, Zhen; Zou, Shuhua

    2016-06-01

    Estradiol mediates its actions by binding to classical nuclear receptors, estrogen receptor α (ER-α) and estrogen receptor β (ER-β), and the non-classical G protein-coupled estrogen receptor 1(GPER). Several gene knockdown models have shown the importance of the receptors for growth of the oocyte and for ovulation. The aim of our study was to identify the pattern of GPER expression in human cumulus granulosa cells (CGCs) from ovarian follicles at different stages of oocyte maturation, and the differences of GPER expression between polycystic ovary syndrome (PCOS) patients and non-PCOS women. Thirty-eight cases of PCOS patients and a control group of thirty-two infertile women without PCOS were used in this study. GPER's location in CGCs was investigated by immunohistochemistry. Quantitative RT-PCR and western blot were used to identify the quantify GPER expression. Here we demonstrated that GPER was expressed in CGCs of both PCOS patients and non-PCOS women, and the expression of GPER was decreased significantly during oocyte maturation. But the expression levels of GPER in CGCs of PCOS patients and non-PCOS women were not significantly different. The data indicate that GPER may play a role during human oocyte maturation through its action in cumulus granulosa cells. AMHRIIs: anti-Mullerian hormone type II receptors; BMI: body mass index; CGCs: cumulus granulosa cells; COH: controlled ovarian hyperstimulation; E2: estradiol; EGFR: epidermal growth factor receptor; ER-α: estrogen receptor; ER-β: estrogen receptor β; FF: follicular fluid; FSH: follicle-stimulating hormone; GCs: granulosa cells; GPER: G protein-coupled estrogen receptor 1; GV: germinal vesicle; GVBD: germinal vesicle breakdown; HCG: human chorionic gonadotropin; IRS: immunoreactive score; IVF-ET: in vitro fertilization and embryo transfer; MI: metaphase I; MII: metaphase II; MAPK: mitogen-activated protein kinase; OCCCs: oocyte corona cumulus complexes; PCOS: polycystic ovarian syndrome; q

  1. 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

  2. Modulation of gene expression in small follicle porcine granulosa cells by human follicle stimulating hormone (hFSH)

    Energy Technology Data Exchange (ETDEWEB)

    Calvo, F.O.; Ryan, R.J.; Woloschak, G.E.

    1986-03-01

    Small follicle (1-3 mm) porcine granulosa cells (SFPGF) were isolated by puncture, aspiration and cultured under standard conditions in DMEM, HEPES, BSA, MIX. At the start of culture, cells were stimulated with 100ng hFSH/ml. At various times afterwards total cellular RNA was prepared using guanidine-hydrochloride solubilization, phenol extraction and precipitation from 3M NaOAc, pH 6.0. RNA was 5'-end labelled with /sup 32/P in a kinase reaction and hybridized to an excess of clone-specific DNA immobilized on nitrocellulose filters using stringent hybridization and wash conditions. After autoradiography the RNA hybridized to the DNA blot filter were quantitated by microdensitometry. Hybridization to parent plasmid was negative. RNA derived from control cultures showed patterns of hybridization similar to those obtained from freshly obtained cells. Results of these experiments demonstrate hFSh induction of RNA specific for transferrin receptor, ..cap alpha..-interferon, H-ras, and K-ras. Increased RNA levels were apparent within 10 min of treatment and had declined by 180 min. Expression of actin, p53 and for RNAs declined by 10 min of hFSH addition but was enhanced by 160 min. Levels of ..beta..-interferon, myc, mos, abl and yb RNAs were not detectable under these conditions. These results demonstrate specific gene modulation in SFPGC cultured with hFSH.

  3. The expression pattern of microRNAs in granulosa cells of subordinate and dominant follicles during the early luteal phase of the bovine estrous cycle.

    Directory of Open Access Journals (Sweden)

    Dessie Salilew-Wondim

    Full Text Available 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.

  4. The four and a half LIM domains 2 (FHL2) regulates ovarian granulosa cell tumor progression via controlling AKT1 transcription

    OpenAIRE

    Hua, G; He, C; Lv, X; Fan, L; Wang, C; Remmenga, S W; Rodabaugh, K J; Yang, L; Lele, S M; Yang, P; Karpf, A R; Davis, J S; Wang, C

    2016-01-01

    The four and a half LIM domains 2 (FHL2) has been shown to play important roles in the regulation of cell proliferation, survival, adhesion, motility and signal transduction in a cell type and tissue-dependent manner. However, the function of FHL2 in ovarian physiology and pathology is unclear. The aim of this study was to determine the role and functional mechanism of FHL2 in the progression of ovarian granulosa cell tumors (GCTs). Immunohistochemical analysis indicated that FHL2 was overexp...

  5. Leucine-enkephalin-like immunoreactivity is localized in luteinizing hormone-producing cells in the axolotl (Ambystoma mexicanum) pituitary.

    Science.gov (United States)

    Suzuki, Hirohumi; Yamamoto, Toshiharu

    2014-02-01

    In this study, we used immunohistochemical techniques to determine the cell type of leucine-enkephalin (Leu-ENK)-immunoreactive cells in the axolotl (Ambystoma mexicanum) pituitary. Immunoreactive cells were scattered throughout the pars distalis except for the dorso-caudal portion. These cells were immuno-positive for luteinizing hormone (LH), but they were immuno-negative for adrenocorticotrophic, growth, and thyroid-stimulating hormones, as well as prolactin. Immunoelectron microscopy demonstrated that Leu-ENK-like substance and LH co-localized within the same secretory granules. Leu-ENK secreted from gonadotrophs may participate in LH secretion in an autocrine fashion, and/or may participate in the release of sex steroids together with LH. Copyright © 2013 Elsevier Ltd. All rights reserved.

  6. Norepinephrine stimulates progesterone production in highly estrogenic bovine granulosa cells cultured under serum-free, chemically defined conditions

    Directory of Open Access Journals (Sweden)

    Piccinato Carla A

    2012-11-01

    Full Text Available Abstract Background Since noradrenergic innervation was described in the ovarian follicle, the actions of the intraovarian catecholaminergic system have been the focus of a variety of studies. We aimed to determine the gonadotropin-independent effects of the catecholamine norepinephrine (NE in the steroid hormone profile of a serum-free granulosa cell (GC culture system in the context of follicular development and dominance. Methods Primary bovine GCs were cultivated in a serum-free, chemically defined culture system supplemented with 0.1% polyvinyl alcohol. The culture features were assessed by hormone measurements and ultrastructural characteristics of GCs. Results GCs produced increasing amounts of estradiol and pregnenolone for 144h and maintained ultrastructural features of healthy steroidogenic cells. Progesterone production was also detected, although it significantly increased only after 96h of culture. There was a highly significant positive correlation between estradiol and pregnenolone production in high E2-producing cultures. The effects of NE were further evaluated in a dose–response study. The highest tested concentration of NE (10 (−7 M resulted in a significant increase in progesterone production, but not in estradiol or pregnenolone production. The specificity of NE effects on progesterone productio n was further investigated by incubating GCs with propranolol (10 (−8 M, a non-selective beta-adrenergic antagonist. Conclusions The present culture system represents a robust model to study the impact of intrafollicular factors, such as catecholamines, in ovarian steroidogenesis and follicular development. The results of noradrenergic effects in the steroidogenesis of GC have implications on physiological follicular fate and on certain pathological ovarian conditions such as cyst formation and anovulation.

  7. Norepinephrine stimulates progesterone production in highly estrogenic bovine granulosa cells cultured under serum-free, chemically defined conditions.

    Science.gov (United States)

    Piccinato, Carla A; Montrezor, Luis H; Collares, Cristhianna A V; Vireque, Alessandra A; Rosa e Silva, Alzira A M

    2012-11-22

    Since noradrenergic innervation was described in the ovarian follicle, the actions of the intraovarian catecholaminergic system have been the focus of a variety of studies. We aimed to determine the gonadotropin-independent effects of the catecholamine norepinephrine (NE) in the steroid hormone profile of a serum-free granulosa cell (GC) culture system in the context of follicular development and dominance. Primary bovine GCs were cultivated in a serum-free, chemically defined culture system supplemented with 0.1% polyvinyl alcohol. The culture features were assessed by hormone measurements and ultrastructural characteristics of GCs. GCs produced increasing amounts of estradiol and pregnenolone for 144h and maintained ultrastructural features of healthy steroidogenic cells. Progesterone production was also detected, although it significantly increased only after 96h of culture. There was a highly significant positive correlation between estradiol and pregnenolone production in high E2-producing cultures. The effects of NE were further evaluated in a dose-response study. The highest tested concentration of NE (10 (-7) M) resulted in a significant increase in progesterone production, but not in estradiol or pregnenolone production. The specificity of NE effects on progesterone production was further investigated by incubating GCs with propranolol (10 (-8) M), a non-selective beta-adrenergic antagonist. The present culture system represents a robust model to study the impact of intrafollicular factors, such as catecholamines, in ovarian steroidogenesis and follicular development. The results of noradrenergic effects in the steroidogenesis of GC have implications on physiological follicular fate and on certain pathological ovarian conditions such as cyst formation and anovulation.

  8. Arsenic induced progesterone production in a caspase-3-dependent manner and changed redox status in preovulatory granulosa cells.

    Science.gov (United States)

    Yuan, Xiao-Hua; Lu, Cai-Ling; Yao, Nan; An, Li-Sha; Yang, Bai-Qing; Zhang, Chuan-Ling; Ma, Xu

    2012-01-01

    Arsenic contamination is a principal environmental health threat throughout the world. However, little is known about the effect of arsenic on steroidogenesis in granulosa cells (GCs). We found that the treatment of preovulatory GCs with arsenite stimulated progesterone production. A significant increase in serum level of progesterone was observed in female Sprague-Dawley rats following arsenite treatment at a dose of 10 mg/L/rat/day for 7 days. Further experiments demonstrated that arsenite treatment did not change the level of intracellular cyclic AMP (cAMP) or phosphorylated ERK1/2 in preovulatory GCs; however, progesterone production was significantly decreased when cAMP-dependent protein kinase (PKA) or ERK1/2 pathway was inhibited. This implied that the effect of arsenite on progesterone production may require cAMP/PKA and ERK1/2 signaling but not depend on them. Furthermore, we found that arsenite decreased intracellular reactive oxygen species (ROS) but increased the antioxidant glutathione (GSH) levels and mitochondrial membrane potential (ΔΨm) in parallel to the changes in progesterone production. Progesterone antagonist blocked the arsenic-stimulated increase of GSH levels. Arsenite treatment induced caspase-3 activation, although no apoptosis was observed. Inhibition of caspase-3 activity significantly decreased progesterone production stimulated by arsenite or follicle-stimulating hormone (FSH). GSH depletion with buthionine sulfoximine led to cell apoptosis in response to arsenite treatment. Collectively, this study demonstrated for the first time that arsenite stimulates progesterone production through cleaved/active caspase-3-dependent pathway, and the increase of GSH level promoted by progesterone production may protect GCs against apoptosis and maintain the steroidogenesis of GCs in response to arsenite treatment. Copyright © 2011 Wiley Periodicals, Inc.

  9. Cryptotanshinone Regulates Androgen Synthesis through the ERK/c-Fos/CYP17 Pathway in Porcine Granulosa Cells

    Directory of Open Access Journals (Sweden)

    Danfeng Ye

    2017-01-01

    Full Text Available The aim of the study is to investigate the molecular mechanism behind androgen reduction in porcine granulosa cells (pGCs with Salvia miltiorrhiza Bunge extract cryptotanshinone. PGCs were isolated from porcine ovaries and identified. Androgen excess model of the pGCs was induced with the MAPK inhibitor PD98059 and then treated with cryptotanshinone. The testosterone level was measured by radioimmunoassay in the culture media. The protein levels of P-ERK1/2, c-Fos, and CYP17 in the cells were measured by western blot. Cryptotanshinone decreased the concentration of testosterone and the protein level of CYP17 and increased the protein levels of P-ERK1/2 and c-Fos in the androgen excess mode. After the c-Fos gene was silenced by infection with c-Fos shRNA lentivirus, we measured the mRNA expression by quantitative RT-PCR and protein level by western blot of P-ERK1/2, c-Fos, and CYP17. This showed that the mRNA expression and protein level of P-ERK1/2 and c-Fos were significantly reduced in the shRNA–c-Fos group compared to the scrambled group, while those of CYP17 were significantly increased. So we concluded that cryptotanshinone can significantly reduce the androgen excess induced by PD98059 in pGCs. The possible molecular mechanism for this activity is regulating the ERK/c-Fos/CYP17 pathway.

  10. LINE1 CpG-DNA Hypomethylation in Granulosa Cells and Blood Leukocytes Is Associated With PCOS and Related Traits.

    Science.gov (United States)

    Sagvekar, Pooja; Mangoli, Vijay; Desai, Sadhana; Patil, Anushree; Mukherjee, Srabani

    2017-04-01

    Altered global DNA methylation is indicative of epigenomic instability concerning chronic diseases. Investigating its incidence and association with polycystic ovary syndrome (PCOS) is essential to understand the etiopathogenesis of this disorder. We assessed global DNA methylation differences in peripheral blood leukocytes (PBLs) and cumulus granulosa cells (CGCs) of controls and women with PCOS; and their association with PCOS and its traits. This study included a total of 102 controls and women with PCOS. Forty-one women undergoing controlled ovarian hyperstimulation (COH) and 61 women not undergoing COH were recruited from in vitro fertilization (IVF) and infertility clinics. DNA methylation was measured by ELISA for 5'-methyl-cytosine content and bisulfite sequencing of 5'-untranslated region (5'-UTR) of long interspersed nucleotide element-1 (LINE1/L1). Total 5'-methyl-cytosine and L1 methylation levels in PBLs and CGCs were similar between controls and women with PCOS. Methylation assessed at CpG sites of L1 5'-UTR revealed a single CpG-site (CpG-4) to be consistently hypomethylated in PBLs of both PCOS groups and CGCs of stimulated PCOS group. In unstimulated women, hypomethylation at CpG-4 was strongly associated with PCOS susceptibility, whereas in stimulated group it showed strong associations with PCOS and its hormonal traits. Furthermore, CGCs demonstrated consistent global and CpG-DNA hypomethylation relative to PBLs, irrespective of normal or disease states. Our study revealed strong association of single hypomethylated CpG-site with PCOS. Identification and characterization of more such methyl-CpG signatures in repetitive elements in larger study populations would provide valuable epigenetic insights into PCOS. Copyright © 2017 by the Endocrine Society

  11. The effect of heat stress on gene expression, synthesis of steroids, and apoptosis in bovine granulosa cells.

    Science.gov (United States)

    Li, Lian; Wu, Jie; Luo, Man; Sun, Yu; Wang, Genlin

    2016-05-01

    Summer heat stress (HS) is a major contributing factor in low fertility in lactating dairy cows in hot environments. Heat stress inhibits ovarian follicular development leading to diminished reproductive efficiency of dairy cows during summer. Ovarian follicle development is a complex process. During follicle development, granulosa cells (GCs) replicate, secrete hormones, and support the growth of the oocyte. To obtain an overview of the effects of heat stress on GCs, digital gene expression profiling was employed to screen and identify differentially expressed genes (DEGs; false discovery rate (FDR) ≤ 0.001, fold change ≥2) of cultured GCs during heat stress. A total of 1211 DEGs including 175 upregulated and 1036 downregulated ones were identified, of which DEGs can be classified into Gene Ontology (GO) categories and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. The results suggested that heat stress triggers a dramatic and complex program of altered gene expression in GCs. We hypothesized that heat stress could induce the apoptosis and dysfunction of GCs. Real-time reverse transcription-polymerase chain reaction (RT-PCR) was used to evaluate the expression of steroidogenic genes (steroidogenic acute regulatory protein (Star), cytochrome P-450 (CYP11A1), CYP19A1, and steroidogenic factor 1 (SF-1)) and apoptosis-related genes (caspase-3, BCL-2, and BAX). Radio immunoassay (RIA) was used to analyze the level of 17β-estradiol (E2) and progesterone (P4). We also assessed the apoptosis of GCs by flow cytometry. Our data suggested that heat stress induced GC apoptosis through the BAX/BCL-2 pathway and reduced the steroidogenic gene messenger RNA (mRNA) expression and E2 synthesis. These results suggest that the decreased function of GCs may cause ovarian dysfunction and offer an improved understanding of the molecular mechanism responsible for the low fertility in cattle in summer.

  12. Exposure of Lactating Dairy Cows to Acute Pre-Ovulatory Heat Stress Affects Granulosa Cell-Specific Gene Expression Profiles in Dominant Follicles

    Science.gov (United States)

    Vanselow, Jens; Vernunft, Andreas; Koczan, Dirk; Spitschak, Marion; Kuhla, Björn

    2016-01-01

    High environmental temperatures induce detrimental effects on various reproductive processes in cattle. According to the predicted global warming the number of days with unfavorable ambient temperatures will further increase. The objective of this study was to investigate effects of acute heat stress during the late pre-ovulatory phase on morphological, physiological and molecular parameters of dominant follicles in cycling cows during lactation. Eight German Holstein cows in established lactation were exposed to heat stress (28°C) or thermoneutral conditions (15°C) with pair-feeding for four days. After hormonal heat induction growth of the respective dominant follicles was monitored by ultrasonography for two days, then an ovulatory GnRH dose was given and follicular steroid hormones and granulosa cell-specific gene expression profiles were determined 23 hrs thereafter. The data showed that the pre-ovulatory growth of dominant follicles and the estradiol, but not the progesterone concentrations tended to be slightly affected. mRNA microarray and hierarchical cluster analysis revealed distinct expression profiles in granulosa cells derived from heat stressed compared to pair-fed animals. Among the 255 affected genes heatstress-, stress- or apoptosis associated genes were not present. But instead, we found up-regulation of genes essentially involved in G-protein coupled signaling pathways, extracellular matrix composition, and several members of the solute carrier family as well as up-regulation of FST encoding follistatin. In summary, the data of the present study show that acute pre-ovulatory heat stress can specifically alter gene expression profiles in granulosa cells, however without inducing stress related genes and pathways and suggestively can impair follicular growth due to affecting the activin-inhibin-follistatin system. PMID:27532452

  13. Adverse eff ects of polymeric nanoparticle poly(ethylene glycol- block-polylactide methyl ether (PEG-b-PLA on steroid hormone secretion by porcine granulosa cells

    Directory of Open Access Journals (Sweden)

    Scsukova Sona

    2017-04-01

    Full Text Available Objectives. Development of nanoparticles (NPs for biomedical applications, including medical imaging and drug delivery, is currently undergoing a dramatic expansion. Diverse effects of different type NPs relating to mammalian reproductive tissues have been demonstrated. Th e objective of this study was to explore the in vitro effects of polymeric nanoparticle poly(ethylene glycol-blockpolylactide methyl ether (PEG-b-PLA NPs on functional state and viability of ovarian granulosa cells (GCs, which play an important role in maintaining ovarian function and female fertility.

  14. Differential gene expression in granulosa cells from polycystic ovary syndrome patients with and without insulin resistance: identification of susceptibility gene sets through network analysis.

    Science.gov (United States)

    Kaur, Surleen; Archer, Kellie J; Devi, M Gouri; Kriplani, Alka; Strauss, Jerome F; Singh, Rita

    2012-10-01

    Polycystic ovary syndrome (PCOS) is a heterogeneous, genetically complex, endocrine disorder of uncertain etiology in women. Our aim was to compare the gene expression profiles in stimulated granulosa cells of PCOS women with and without insulin resistance vs. matched controls. This study included 12 normal ovulatory women (controls), 12 women with PCOS without evidence for insulin resistance (PCOS non-IR), and 16 women with insulin resistance (PCOS-IR) undergoing in vitro fertilization. Granulosa cell gene expression profiling was accomplished using Affymetrix Human Genome-U133 arrays. Differentially expressed genes were classified according to gene ontology using ingenuity pathway analysis tools. Microarray results for selected genes were confirmed by real-time quantitative PCR. A total of 211 genes were differentially expressed in PCOS non-IR and PCOS-IR granulosa cells (fold change≥1.5; P≤0.001) vs. matched controls. Diabetes mellitus and inflammation genes were significantly increased in PCOS-IR patients. Real-time quantitative PCR confirmed higher expression of NCF2 (2.13-fold), TCF7L2 (1.92-fold), and SERPINA1 (5.35-fold). Increased expression of inflammation genes ITGAX (3.68-fold) and TAB2 (1.86-fold) was confirmed in PCOS non-IR. Different cardiometabolic disease genes were differentially expressed in the two groups. Decreased expression of CAV1 (-3.58-fold) in PCOS non-IR and SPARC (-1.88-fold) in PCOS-IR was confirmed. Differential expression of genes involved in TGF-β signaling (IGF2R, increased; and HAS2, decreased), and oxidative stress (TXNIP, increased) was confirmed in both groups. Microarray analysis demonstrated differential expression of genes linked to diabetes mellitus, inflammation, cardiovascular diseases, and infertility in the granulosa cells of PCOS women with and without insulin resistance. Because these dysregulated genes are also involved in oxidative stress, lipid metabolism, and insulin signaling, we hypothesize that these

  15. Circular RNA expression profiling of human granulosa cells during maternal aging reveals novel transcripts associated with assisted reproductive technology outcomes.

    Directory of Open Access Journals (Sweden)

    Jing Cheng

    Full Text Available Circular RNAs (circRNAs are a unique class of endogenous RNAs which could be used as potential diagnostic and prognostic biomarkers of many diseases. Our study aimed to investigate circRNA profiles in human granulosa cells (GCs during maternal aging and to uncover age-related circRNA variations that potentially reflect decreased oocyte competence. CircRNAs in GCs from in vitro fertilization (IVF patients with young age (YA, ≤ 30 years and advanced age (AA, ≥ 38 years were profiled by microarray, and validated in 20 paired samples. The correlation between circRNAs expression and clinical characteristics was analyzed in additional 80 samples. Chip-based analysis revealed 46 up-regulated and 11 down-regulated circRNAs in AA samples (fold change > 2.0. Specifically, circRNA_103829, circRNA_103827 and circRNA_104816 were validated to be up-regulated, while circRNA_101889 was down-regulated in AA samples. After adjustment for gonadotropin treatment, only circRNA_103827 and circRNA_104816 levels were positively associated with maternal age (partial r = 0.332, P = 0.045; partial r = 0.473, P = 0.003; respectively. Moreover, circRNA_103827 and circRNA_104816 expressions in GCs were negatively correlated with the number of top quality embryos (r = -0.235, P = 0.036; r = -0.221, P = 0.049; respectively. Receiver operating characteristic (ROC curve analysis indicated that the performance of circRNA_103827 for live birth prediction reached 0.698 [0.570-0.825], with 77.2% sensitivity and 60.9% specificity (P = 0.006, and that of circRNA_104816 was 0.645 [0.507-0.783] (P = 0.043. Bioinformatics analysis revealed that both circRNAs were potentially involved in glucose metabolism, mitotic cell cycle, and ovarian steroidogenesis. Therefore, age-related up-regulation of circRNA_103827 and circRNA_104816 might be potential indicators of compromised follicular micro-environment which could be used to predict IVF prognosis, and improve female infertility

  16. Luteinizing hormone-induced Akt phosphorylation and androgen production are modulated by MAP Kinase in bovine theca cells

    Directory of Open Access Journals (Sweden)

    Fukuda Shin

    2009-11-01

    Full Text Available Abstract Background Theca cells play an important role in controlling ovarian steroidogenesis by providing aromatizable androgens for granulosa cell estrogen biosynthesis. Although it is well established that the steroidogenic activity of theca cells is mainly regulated by LH, the intracellular signal transduction mechanisms that regulate thecal proliferation and/or steroidogenesis remain obscure. In this study, we examined whether and how LH controls the PI3K/Akt signaling pathway and androgen production in bovine theca cells. We also explored whether this LH-induced PI3K/Akt activation is modulated with other signaling pathways (i.e. PKA and MAPK. Methods Ovarian theca cells were isolated from bovine small antral follicles and were incubated with LH for various durations. Phospho-Akt and total-Akt content in the cultured theca cells were examined using Western blotting. Androstenedione levels in the spent media were determined using EIA. Semi-quantitative RT-PCR analyses were conducted to analyze the mRNA levels of CYP17A1 and StAR in the theca cells. To examine whether Akt activity is involved in theca cell androgen production, the PI3K inhibitors wortmannin and LY294002 were also added to the cells. Results Akt is constitutively expressed, but is gradually phosphorylated in cultured bovine theca cells through exposure to LH. LH significantly increased androstenedione production in bovine theca cells, whereas addition of the wortmannin and LY294002 significantly decreased LH-induced androstenedione production. LH significantly increased CYP17A1 mRNA level in theca cells, whereas addition of LY294002 significantly decreased LH-induced CYP17A1 expression. Neither LH nor PI3K inhibitors alter the mRNA levels of StAR in theca cells. Although H89 (a selective inhibitor of PKA does not affect LH-mediated changes in Akt, U0126 (a potent MEK inhibitor suppressed LH-induced Akt phosphorylation, CYP17A1 expression, and androgen production in theca

  17. Testosterone-Dependent Interaction between Androgen Receptor and Aryl Hydrocarbon Receptor Induces Liver Receptor Homolog 1 Expression in Rat Granulosa Cells

    Science.gov (United States)

    Wu, Yanguang; Baumgarten, Sarah C.; Zhou, Ping

    2013-01-01

    Androgens play a major role in the regulation of normal ovarian function; however, they are also involved in the development of ovarian pathologies. These contrasting effects may involve a differential response of granulosa cells to the androgens testosterone (T) and dihydrotestosterone (DHT). To determine the molecular pathways that mediate the distinct effects of T and DHT, we studied the expression of the liver receptor homolog 1 (LRH-1) gene, which is differentially regulated by these steroids. We found that although both T and DHT stimulate androgen receptor (AR) binding to the LRH-1 promoter, DHT prevents T-mediated stimulation of LRH-1 expression. T stimulated the expression of aryl hydrocarbon receptor (AHR) and its interaction with the AR. T also promoted the recruitment of the AR/AHR complex to the LRH-1 promoter. These effects were not mimicked by DHT. We also observed that the activation of extracellular regulated kinases by T is required for AR and AHR interaction. In summary, T, but not DHT, stimulates AHR expression and the interaction between AHR and AR, leading to the stimulation of LRH-1 expression. These findings could explain the distinct response of granulosa cells to T and DHT and provide a molecular mechanism by which DHT negatively affects ovarian function. PMID:23689136

  18. Hydrostatic pressure affects in vitro maturation of oocytes and follicles and increases granulosa cell death.

    Science.gov (United States)

    Rashidi, Zahra; Azadbakht, Mehri; Amini, Ali; Karimi, Isac

    2014-01-01

    This study examines the effects of hydrostatic pressure on in vitro maturation (IVM) of oocytes derived from in vitro grown follicles. In this experimental study, preantral follicles were isolated from 12-day-old female NMRI mice. Each follicle was cultured individually in Alpha Minimal Essential Medium (α-MEM) under mineral oil for 12 days. Then, follicles were induced for IVM and divided into two groups, control and experiment. In the experiment group follicles were subjected to 20 mmHg pressure for 30 minutes and cultured for 24-48 hours. We assessed for viability and IVM of the oocytes. The percentage of apoptosis in cumulus cells was determined by the TUNEL assay. A comparison between groups was made using the student's t test. The percentage of metaphase II oocytes (MII) increased in hydrostatic pressuretreated follicles compared to controls (phydrostatic pressure-treated follicles compared to controls (pHydrostatic pressure, by inducing apoptosis in cumulus cells, participates in the cumulus oocyte coupled relationship with oocyte maturation.

  19. The incidence of endometrial hyperplasia and cancer in 1031 patients with a granulosa cell tumor of the ovary: long-term follow-up in a population-based cohort study

    NARCIS (Netherlands)

    van Meurs, Hannah S.; Bleeker, Maaike C. G.; van der Velden, Jacobus; Overbeek, Lucy I. H.; Kenter, Gemma G.; Buist, Marrije R.

    2013-01-01

    Concurrent presence of endometrial hyperplasia or cancer in patients with granulosa cell tumors (GCTs) is common, with reported incidences of 25.6% to 65.5%. Consequently, bilateral salpingo-oophorectomy and hysterectomy is usually recommended in patients with a GCT, but this remains debatable. Our

  20. MicroRNA Expression Profile in Bovine Granulosa Cells of Preovulatory Dominant and Subordinate Follicles during the Late Follicular Phase of the Estrous Cycle.

    Directory of Open Access Journals (Sweden)

    Samuel Gebremedhn

    Full Text Available In bovine, ovarian follicles grow in a wave-like fashion with commonly 2 or 3 follicular waves emerging per estrous cycle. The dominant follicle of the follicular wave which coincides with the LH-surge becomes ovulatory, leaving the subordinate follicles to undergo atresia. These physiological processes are controlled by timely and spatially expressed genes and gene products, which in turn are regulated by post-transcriptional regulators. MicroRNAs, a class of short non-coding RNA molecules, are one of the important posttranscriptional regulators of genes associated with various cellular processes. Here we investigated the expression pattern of miRNAs in granulosa cells of bovine preovulatory dominant and subordinate follicles during the late follicular phase of bovine estrous cycle using Illumina miRNA deep sequencing. In addition to 11 putative novel miRNAs, a total of 315 and 323 known miRNAs were detected in preovulatory dominant and subordinate follicles, respectively. Moreover, in comparison with the subordinate follicles, a total of 64 miRNAs were found to be differentially expressed in preovulatory dominant follicles, of which 34 miRNAs including the miR-132 and miR-183 clusters were significantly enriched, and 30 miRNAs including the miR-17-92 cluster, bta-miR-409a and bta-miR-378 were significantly down regulated in preovulatory dominant follicles. In-silico pathway analysis revealed that canonical pathways related to oncogenesis, cell adhesion, cell proliferation, apoptosis and metabolism were significantly enriched by the predicted target genes of differentially expressed miRNAs. Furthermore, Luciferase reporter assay analysis showed that one of the differentially regulated miRNAs, the miR-183 cluster miRNAs, were validated to target the 3'-UTR of FOXO1 gene. Moreover FOXO1 was highly enriched in granulosa cells of subordinate follicles in comparison with the preovulatory dominant follicles demonstrating reciprocal expression pattern

  1. Efficient differentiation of steroidogenic and germ-like cells from epigenetically-related iPSCs derived from ovarian granulosa cells.

    Directory of Open Access Journals (Sweden)

    Raymond Anchan

    Full Text Available To explore restoration of ovarian function using epigenetically-related, induced pluripotent stem cells (iPSCs, we functionally evaluated the epigenetic memory of novel iPSC lines, derived from mouse and human ovarian granulosa cells (GCs using c-Myc, Klf4, Sox2 and Oct4 retroviral vectors. The stem cell identity of the mouse and human GC-derived iPSCs (mGriPSCs, hGriPSCs was verified by demonstrating embryonic stem cell (ESC antigen expression using immunocytochemistry and RT-PCR analysis, as well as formation of embryoid bodies (EBs and teratomas that are capable of differentiating into cells from all three germ layers. GriPSCs' gene expression profiles associate more closely with those of ESCs than of the originating GCs as demonstrated by genome-wide analysis of mRNA and microRNA. A comparative analysis of EBs generated from three different mouse cell lines (mGriPSCs; fibroblast-derived iPSC, mFiPSCs; G4 embryonic stem cells, G4 mESCs revealed that differentiated mGriPSC-EBs synthesize 10-fold more estradiol (E2 than either differentiated FiPSC- or mESC-EBs under identical culture conditions. By contrast, mESC-EBs primarily synthesize progesterone (P4 and FiPSC-EBs produce neither E2 nor P4. Differentiated mGriPSC-EBs also express ovarian markers (AMHR, FSHR, Cyp19a1, ER and Inha as well as markers of early gametogenesis (Mvh, Dazl, Gdf9, Boule and Zp1 more frequently than EBs of the other cell lines. These results provide evidence of preferential homotypic differentiation of mGriPSCs into ovarian cell types. Collectively, our data support the hypothesis that generating iPSCs from the desired tissue type may prove advantageous due to the iPSCs' epigenetic memory.

  2. Lutein and Brain Function

    Directory of Open Access Journals (Sweden)

    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.

  3. Bilateral granulosa cell tumors: a novel malignant manifestation of multiple endocrine neoplasia 1 syndrome found in a patient with a rare menin in-frame deletion

    Directory of Open Access Journals (Sweden)

    Hall MJ

    2015-02-01

    Full Text Available Michael J Hall,1 Julie Innocent,2 Christina Rybak,1 Colleen Veloski,3 Walter J Scott,4 Hong Wu,5 John A Ridge,4 John P Hoffman,4 Hossein Borghaei,2 Aruna Turaka,6 Mary B Daly1 1Department of Clinical Genetics, 2Department of Medical Oncology, 3Department of Internal Medicine, 4Department of Surgical Oncology, 5Department of Pathology, 6Department of Radiation Oncology, Fox Chase Cancer Center, Philadelphia, PA, USA Introduction: Multiple endocrine neoplasia 1 (MEN1 is a cancer syndrome resulting from mutations of the MEN1 gene. The syndrome is characterized by neoplasia of the parathyroid and pituitary glands, and malignant tumors of the endocrine pancreas. Other manifestations include benign lipomas, angiofibromas, and carcinoid tumors commonly originating in the colon, thymus, and lung. This is the first report of MEN1 syndrome manifesting as bilateral granulosa cell ovarian tumors, and which is associated with a rare intronic mutation of the MEN1 gene. Case report: A 41-year-old woman presented with abdominal pain, increasing abdominal girth, and dysmenorrhea. Ultrasound demonstrated enlarged ovaries and uterine fibroids. After an exploratory laparotomy, she subsequently underwent bilateral salpingo–oophorectomy with hysterectomy where the pathology revealed bilateral cystic granulosa cell tumors of the ovaries. Additional workup including computed tomography imaging discovered a thymic mass, which the pathology showed was malignant, along with a pancreatic mass suspicious for a neuroendocrine tumor. Hyperparathyroidism was also discovered and was found to be secondary to a parathyroid adenoma. Genetic testing revealed an exceedingly rare mutation in the MEN1 gene (c.654 + 1 G>A. Discussion: Mutations of the menin gene leading to MEN1 syndrome are classically nonsense or missense mutations producing a dysfunctional protein product. Recently, researchers described a novel mutation of MEN1 (c.654 + 1 G>A in a male proband meeting the criteria

  4. Protective role of melatonin in progesterone production by human luteal cells.

    Science.gov (United States)

    Taketani, Toshiaki; Tamura, Hiroshi; Takasaki, Akihisa; Lee, Lifa; Kizuka, Fumie; Tamura, Isao; Taniguchi, Ken; Maekawa, Ryo; Asada, Hiromi; Shimamura, Katsunori; Reiter, Russel J; Sugino, Norihiro

    2011-09-01

    This study investigated whether melatonin protects luteinized granulosa cells from reactive oxygen species (ROS) as an antioxidant to enhance progesterone production in the follicle during ovulation. Follicular fluid was sampled at the time of oocyte retrieval in women undergoing in vitro fertilization and embryo transfer (IVF-ET). Melatonin concentrations in the follicular fluid were positively correlated with progesterone concentrations (r = 0.342, P progesterone and 8-OHdG concentrations were negatively correlated (r = -0.246, P Progesterone production by luteinized granulosa cells was significantly inhibited by H(2)O(2). Melatonin treatment overcame the inhibitory effect of H(2) O(2) . Twenty-five patients who had luteal phase defect (serum progesterone concentrations progesterone concentrations (>10 ng/mL during the mid-luteal phase) in nine of 14 women (64.3%), whereas only two of 11 women (18.1%) showed normal serum progesterone levels in the control group. In conclusion, melatonin protects granulosa cells undergoing luteinization from ROS in the follicle and contributes to luteinization for progesterone production during ovulation. © 2011 John Wiley & Sons A/S.

  5. Follicle-stimulating hormone (FSH) unmasks specific high affinity FSH-binding sites in cell-free membrane preparations of porcine granulosa cells

    Energy Technology Data Exchange (ETDEWEB)

    Ford, K.A.; LaBarbera, A.R.

    1988-11-01

    The purpose of these studies was to determine whether changes in FSH receptors correlated with FSH-induced attenuation of FSH-responsive adenylyl cyclase in immature porcine granulosa cells. Cells were incubated with FSH (1-1000 ng/ml) for up to 24 h, treated with acidified medium (pH 3.5) to remove FSH bound to cells, and incubated with (125I)iodo-porcine FSH to quantify FSH-binding sites. FSH increased binding of FSH in a time-, temperature-, and FSH concentration-dependent manner. FSH (200 ng/ml) increased binding approximately 4-fold within 16 h. Analysis of equilibrium saturation binding data indicated that the increase in binding sites reflected a 2.3-fold increase in receptor number and a 5.4-fold increase in apparent affinity. The increase in binding did not appear to be due to 1) a decrease in receptor turnover, since the basal rate of turnover appeared to be very slow; 2) an increase in receptor synthesis, since agents that inhibit protein synthesis and glycosylation did not block the increase in binding; or 3) an increase in intracellular receptors, since agents that inhibit cytoskeletal components had no effect. Agents that increase intracellular cAMP did not affect FSH binding. The increase in binding appeared to result from unmasking of cryptic FSH-binding sites, since FSH increased binding in cell-free membrane preparations to the same extent as in cells. Unmasking of cryptic sites was hormone specific, and the sites bound FSH specifically. Unmasking of sites was reversible in a time- and temperature-dependent manner after removal of bound FSH. The similarity between the FSH dose-response relationships for unmasking of FSH-binding sites and attenuation of FSH-responsive cAMP production suggests that the two processes are functionally linked.

  6. Nearly 30 Years of Treatment for Recurrent Granulosa Cell Tumor of the Ovary: A Case Report and Review of the Literature

    Directory of Open Access Journals (Sweden)

    Deanna Teoh

    2010-01-01

    Full Text Available A 30-year-old woman was diagnosed with a stage IA granulosa cell tumor (GCT of the ovary in 1979. Following removal of the adnexal mass and complete surgical staging, she remained disease-free for 12 years. In 1991 she underwent a resection of a retroperitoneal mass, confirmed to be a recurrent GCT. Despite adjuvant radiation treatment at the time of recurrence, the patient presented five years later with abdominal pain, and was found to have a second recurrence. Over the next 10 years the patient had multiple recurrences and progressive disease despite surgical resection, cytotoxic, hormonal and targeted chemotherapy treatments. In conclusion, there is no standard management for recurrent GCT of the ovary. We review this patient’s treatment in the context of the current literature.

  7. Concentration of activin A and follistatin in follicular fluid from human small antral follicles associated to gene expression of the corresponding granulosa cells

    DEFF Research Database (Denmark)

    Jeppesen, J V; Nielsen, M E; Kristensen, S G

    2012-01-01

    The present study correlated concentrations of activin A and follistatin in follicular fluid (FF) from human small antral follicles to FF concentrations of AMH, inhibin B, progesterone, and oestradiol and to the mRNA expression of FSH-receptor (FSHR), LH-receptor (LHR), AMH-receptor2 (AMHR2), CYP19...... activin A levels increased in follicles exceeding 10 mm in diameter. Levels of activin A and inhibin B showed a highly significant inverse association. Follistatin showed highly significant positive associations with AMH and inhibin B levels and with FSHR and AR gene expression in GC. This study revealed......a, and androgen-receptor (AR) in the corresponding granulosa cells (GC). FF from 144 follicles (3-12 mm in diameter) was included whereas mRNA expression profiles were established in GC from 66 of the 144 follicles. Levels of follistatin remained constant in relation to follicular diameter, whereas...

  8. Hexavalent chromium-induced apoptosis of granulosa cells involves selective sub-cellular translocation of Bcl-2 members, ERK1/2 and p53

    International Nuclear Information System (INIS)

    Banu, Sakhila K.; Stanley, Jone A.; Lee, JeHoon; Stephen, Sam D.; Arosh, Joe A.; Hoyer, Patricia B.; Burghardt, Robert C.

    2011-01-01

    Hexavalent chromium (CrVI) has been widely used in industries throughout the world. Increased usage of CrVI and atmospheric emission of CrVI from catalytic converters of automobiles, and its improper disposal causes various health hazards including female infertility. Recently we have reported that lactational exposure to CrVI induced a delay/arrest in follicular development at the secondary follicular stage. In order to investigate the underlying mechanism, primary cultures of rat granulosa cells were treated with 10 μM potassium dichromate (CrVI) for 12 and 24 h, with or without vitamin C pre-treatment for 24 h. The effects of CrVI on intrinsic apoptotic pathway(s) were investigated. Our data indicated that CrVI: (i) induced DNA fragmentation and increased apoptosis, (ii) increased cytochrome c release from the mitochondria to cytosol, (iii) downregulated anti-apoptotic Bcl-2, Bcl-XL, HSP70 and HSP90; upregulated pro-apoptotic BAX and BAD, (iv) altered translocation of Bcl-2, Bcl-XL, BAX, BAD, HSP70 and HSP90 to the mitochondria, (v) upregulated p-ERK and p-JNK, and selectively translocated p-ERK to the mitochondria and nucleus, (vi) activated caspase-3 and PARP, and (vii) increased phosphorylation of p53 at ser-6, ser-9, ser-15, ser-20, ser-37, ser-46 and ser-392, increased p53 transcriptional activation, and downregulated MDM-2. Vitamin C pre-treatment mitigated CrVI effects on apoptosis and related pathways. Our study, for the first time provides a clear insight into the effect of CrVI on multiple pathways that lead to apoptosis of granulosa cells which could be mitigated by vitamin C.

  9. Morphometric analysis of the folliculostellate cells and luteinizing hormone gonadotropic cells of the anterior pituitary of the men during the aging process.

    Science.gov (United States)

    Čukuranović Kokoris, Jovana; Jovanović, Ivan; Pantović, Vukica; Krstić, Miljan; Stanojković, Milica; Milošević, Verica; Ugrenović, Slađana; Stojanović, Vesna

    2017-02-01

    The aim of this research was to quantify the changes in the morphology and density of the anterior pituitary folliculostellate (FS) and luteinizing hormone (LH) cells. Material was tissue of the pituitary gland of the 14 male cadavers. Tissue slices were immunohistochemically stained with monoclonal anti-LH antibody and polyclonal anti-S100 antibody for the detection of LH and FS cells, respectively. Digital images of the stained slices were afterwards morphometrically analyzed by ImageJ. Results of the morphometric analysis showed significant increase of the FS cells volume density in cases older than 70 years. Volume density of the LH cells did not significantly change, whereas their area significantly increased with age. Nucleocytoplasmic ratio of the LH cells gradually decreased and became significant after the age of 70. Finally, volume density of the FS cell significantly correlated with LH cells area and nucleocytoplasmic ratio. From all above cited, we concluded that in men, density and size of the FS cells increase with age. Long-term hypertrophy of the LH cells results in their functional decline after the age of 70. Strong correlation between FS cells and LH cells morphometric parameters might point to age-related interaction between these two cell groups. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Influence of ionizing radiation, photoperiod, and environmental temperature on cell proliferation in the intestinal epithelium of the rough-skinned newt (Taricha granulosa)

    International Nuclear Information System (INIS)

    Filipy, R.E.

    1977-01-01

    Kinetics of cell division in the intestinal epithelial proliferative cells (cell nests) of the rough-skinned newt, Taricha granulosa, were studied using tritiated thymidine autoradiography and the mitotic arresting properties of colcemid. Percent labeled mitoses (PLM) curves were drawn from the autoradiographic data from two separate experiments in which the newts were maintained at room temperature (22 to 23 0 C). In those experiments, groups of newts were intraperitoneally injected with tritiated thymidine at times ranging between 5 and 58 hours before they were killed. Each newt in both experiments was intraperitoneally injected with colcemid five hours before it was killed. Data from both experiments were very similar and the PLM curves were used to estimate cell cycle phase durations in the cell nests. The DNA synthetic phase was estimated to be 41 hours long and the sum of the G 2 phase duration and one-half the duration of mitosis was approximately seven hours. The duration of the entire cell cycle was considered to be longer than 100 hours and the duration of the G 1 phase longer than 50 hours. Whole-body exposure of newts to 100 R x irradiation essentially stopped mitotic activity in the intestinal cell nests for 32 hours and exposure to 200 R stopped the mitotic activity for 56 hours. Recovery of mitotic activity in the cell nests was in evidence in data from both groups of animals after both those time periods. Whole-body exposure of newts to 1000 R x irradiation severely depressed mitotic activity in the intestinal cell nests for at least 78 hours and resulted in diminished numbers of the cell nests although the mature cells of the intestinal cells appeared to be relatively radioresistant

  11. Opposite Effects of the Spinach Food Matrix on Lutein Bioaccessibility and Intestinal Uptake Lead to Unchanged Bioavailability Compared to Pure Lutein.

    Science.gov (United States)

    Margier, Marielle; Buffière, Caroline; Goupy, Pascale; Remond, Didier; Halimi, Charlotte; Caris-Veyrat, Catherine; Borel, Patrick; Reboul, Emmanuelle

    2018-06-01

    Food matrix is generally believed to alter carotenoid bioavailability, but its effect on xanthophylls is usually limited. This study thus aims to decipher the digestion-absorption process of lutein in the presence or not of a food matrix. Lutein transfer to gastric-like lipid droplets or artificial mixed micelles was assessed when lutein was added to test meals either as a pure molecule ((all-E)-lutein) or in canned spinach ((Z) + (all-E)-lutein). The obtained mixed micelles were delivered to Caco-2 cells to evaluate lutein uptake. Finally postprandial plasma lutein responses were compared in minipigs after the two test meals. Lutein transfer to gastric-like lipid droplets and to mixed micelles was higher when lutein was added in spinach than when it was added as pure lutein (+614% and +147%, respectively, p < 0.05). Conversely, lutein uptake was less effective when micellar lutein was from a meal containing spinach than from a meal containing its pure form (-55%, p < 0.05). In minipigs, postprandial lutein response was delayed with spinach but not significantly different after the two test meals. Opposite effects at the micellarization and intestinal cell uptake steps explain the lack of effect of spinach matrix on lutein bioavailability. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. 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.

  13. Molecular cloning and expression analyses of porcine MAP1LC3A in the granulosa cells of normal and miniature pig

    Directory of Open Access Journals (Sweden)

    Kim Sang H

    2013-02-01

    Full Text Available Abstract Background The members of the microtubule-associated protein 1 light chain (MAP1LC family, especially those of the LC3 family (MAP1LC3A, B, C, are known to induce autophagy upon localization onto the autophagosomal membrane. In this regard, LC3 can be utilized as a marker for the formation of autophagosomes during the process of autophagy. The aims of this study are to clone porcine MAP1LC3A, and analyze the pattern of its expression in the ovarian tissues of normal and miniature pig ovary in an attempt to understand the distinct mode of apoptosis between two strains. Methods Rapid amplification of cDNA ends (RACE were used to obtain the 5′ and 3′ ends of the porcine MAP1LC3A full length cDNA. Reverse-transcriptase-PCR (RT-PCR, real-time PCR, and western blot analysis were performed to examine the expression of porcine MAP1LC3A. The localization of MAP1LC3A in the ovary was determined by In situ Hybridization and Immunohistochemical staining. Results We cloned the full-length cDNA of porcine MAP1LC3A and identified an open reading frame of 980 bp encoding 121 amino acids. Based on its homology to known mammalian proteins (98% this novel cDNA was designated as porcine MAP1LC3A and registered to the GenBank (Accession No. GU272221. We compared the expression of MAP1LC3A in the Graafian follicles of normal and miniature pigs by in situ hybridization at day 15 of the estrus cycle. While normal pigs showed a stronger expression of MAP1LC3A mRNA than miniature pigs in the theca cell area, the expression was lower in the granulosa cells. Immunofluorescence analysis of the MAP1LC3A fusion reporter protein showed the subcellular localization of porcine MAP1LC3A and ATG5 as a punctate pattern in the cytoplasm of porcine granulosa cells under stress conditions. In addition, the expressions of MAP1LC3A and ATG5 were higher in normal pigs than in miniature pigs both in the presence and absence of rapamycin. Conclusions The newly cloned porcine

  14. A positive feedback loop between progesterone and microsomal prostaglandin E synthase-1-mediated PGE2 promotes production of both in mouse granulosa cells.

    Science.gov (United States)

    Tamura, Kazuhiro; Naraba, Hiroaki; Hara, Takahiko; Nakamura, Kota; Yoshie, Mikihiro; Kogo, Hiroshi; Tachikawa, Eiichi

    2016-03-01

    Microsomal prostaglandin E synthase-1 (mPGES-1) is primarily expressed in granulosa cells (GCs) in the preovulatory follicle. Both prostaglandin E2 (PGE2) and progesterone (P4) are implicated in various reproductive functions. Here, we demonstrate that mPges-1 may be a direct downstream target gene of the P4 receptor and P4-stimulated PGE2 secretion can stimulate P4 production in a newly generated mouse GC line (GtsT). Treatment of GtsT cells with a P4 receptor agonist, norgestrel, markedly increased mPGES-1 expression detected by RT-PCR analysis. PGE2 secretion measured by an enzyme-linked immunosorbent assay was enhanced by P4 treatment. Luciferase assays revealed that the proximal promoter region of the mPges-1 gene was responsible for the effects of P4 treatment. Conversely, PGE2 treatment stimulated P4 secretion, which coordinated with mRNA expression of steroidogenic acute regulatory protein. Taken together, P4 may regulate mPGES-1 expression to increase PGE2 secretion and in turn P4 production. An autocrine loop between P4 and PGE2 might function to maintain the increased levels of both in GCs. Copyright © 2016 Elsevier Inc. All rights reserved.

  15. ANP promotes proliferation and inhibits apoptosis of ovarian granulosa cells by NPRA/PGRMC1/EGFR complex and improves ovary functions of PCOS rats.

    Science.gov (United States)

    Zheng, Qin; Li, Yulin; Zhang, Dandan; Cui, Xinyuan; Dai, Kuixing; Yang, Yu; Liu, Shuai; Tan, Jichun; Yan, Qiu

    2017-10-26

    Polycystic ovary syndrome (PCOS) is a complicated reproductive endocrine disease characterized by polycystic ovaries, hyperandrogenism and anovulation. It is one of the main causes of infertility. RU486 is an antagonist of progesterone receptor, and most commonly used as a contraceptive. However, whether RU486 is correlated with PCOS remains unclear. Atrial natriuretic peptide (ANP) is a small peptide with natriuretic and diuretic functions, and its availability to be used in PCOS treatment is unknown. Here, we showed that the serum ANP level was lower in PCOS patients than that in healthy women, and it was also decreased in the serum and ovarian tissues of RU486-induced PCOS rats compared with the control rats. We also found that RU486 inhibited the proliferation and promoted the apoptosis of human KGN ovarian granulosa cells by downregulating progesterone receptor membrane component 1 (PGRMC1). Meantime, ANP promoted the proliferation and inhibited the apoptosis of KGN cells through upregulating ANP receptor A (NPRA). The promotive effects of ANP on ovarian functions were mediated through the formation of an NPRA/PGRMC1/EGFR complex, which further activated MAPK/ERK signaling and transcription factor AP1. Moreover, ANP treatment reversed the PCOS symptoms, and improved the fertility of RU486-induced PCOS rats. Collectively, these findings highlight that RU486 is associated with the pathogenesis of PCOS, and ANP treatment may be a promising therapeutic option for PCOS.

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

    Directory of Open Access Journals (Sweden)

    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. Keywords: IRS-2, TFBS, FSH, SP1, ChIP

  17. LncRNA-LET inhibits cell viability, migration and EMT while induces apoptosis by up-regulation of TIMP2 in human granulosa-like tumor cell line KGN.

    Science.gov (United States)

    Han, Qingfang; Zhang, Wenke; Meng, Jinlai; Ma, Li; Li, Aihua

    2018-04-01

    Polycystic ovary syndrome (PCOS) is a common endocrine disease characterized by hyperandrogenism, irregular menses, and polycystic ovaries. Several long non-coding RNAs (lncRNAs) are aberrantly expressed in PCOS patients; however, little is known about the effects of the lncRNA-low expression in tumor (lncRNA-LET) on PCOS. We aimed to explore the effects of lncRNA-LET on human granulosa-like tumor cell line, KGN. Expression of lncRNA-LET in normal IOSE80 cells and granulosa cells was determined by qRT-PCR. KGN cell viability, apoptosis and migration were measured by trypan blue exclusion method, flow cytometry assay and wound healing assay, respectively. TGF-β1 was used to induce epithelial-mesenchymal transition (EMT) process. LncRNA-LET expression and mRNA expressions of TIMP2 and EMT-related proteins were measured by qRT-PCR. Western blot analysis was used to measure the protein expression of apoptosis-related proteins, EMT-related proteins, TIMP2, and the proteins in the Wnt/β-catenin and Notch signaling pathways. lncRNA-LET was down-regulated in KGN cells, and its overexpression inhibited cell viability and migration, and promoted apoptosis in KGN cells. Overexpression of lncRNA-LET increased the expression of E-cadherin and decreased the expressions of N-cadherin and vimentin in KGN cells. These effects of lncRNA-LET on KGN cells were reversed by TIMP2 suppression. Overexpression of TIMP2 inhibited cell viability, migration and EMT process, and increased apoptosis by activating the Wnt/β-catenin and Notch pathways. Overexpression of lncRNA-LET inhibits cell viability, migration and EMT process, and increases apoptosis in KGN cells by up-regulating the expression of TIMP2 and activating the Wnt/β-catenin and notch signaling pathways. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

  18. Adult Type Granulosa Cell Tumor: A Very Rare Case of Sex-Cord Tumor of the Testis with Review of the Literature

    Directory of Open Access Journals (Sweden)

    Dimosthenis Miliaras

    2013-01-01

    Full Text Available Granulosa cell tumor (GST is a sex-cord/stromal neoplasm of the gonads, more commonly arising in the ovaries, while approximately 80 cases have been reported in the testes. Out of these, 30 cases were of the adult type, while the remainder 50 cases were of the juvenile type. The latter mostly concerned infants and followed a benign course. However, the adult type testicular GCTs may be potentially malignant as it also happens in female patients with such neoplasms. We present a case of an adult type GCT located at the left testis. The patient was subjected to total orchiectomy and received no further treatment. Histology showed typical GCT histomorphology with Call-Exner bodies in some places. The immunoprofile of the tumor was CD99 (+, calretinin (+, inhibin (+, alpha smooth muscle actin (+, vimentin (+, ER (−, PR (−, keratin AE1/AE3 (−, alpha fetoprotein (−, CD117 (−, and placental alkaline phosphatase (−. Two years after surgery, the patient is alive and well with no signs of recurrence.

  19. Effectiveness of different treatment modalities for the management of adult-onset granulosa cell tumours of the ovary (primary and recurrent).

    Science.gov (United States)

    Gurumurthy, Mahalakshmi; Bryant, Andrew; Shanbhag, Smruta

    2014-04-21

    Granulosa cell tumour is a rare gynaecological tumour of the ovary with recurrences many years after initial diagnosis and treatment. Evidence-based management of granulosa cell tumour of the ovary is limited, and treatment has not been standardised. Surgery, including fertility-sparing procedures for young women, has traditionally been the standard treatment. Adjuvant treatments following surgery have been based on non-randomised trials. A combination of bleomycin, etoposide and cisplatin (BEP) has traditionally been used for treatment of advanced and/or recurrent disease that cannot be optimally managed surgically. To evaluate the effectiveness and safety of different treatment modalities offered in current practice for the management of primary, residual and recurrent adult-onset granulosa cell tumours (GCTs) of the ovary. We searched the Cochrane Gynaecological Cancer Group Trials Register, the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE and EMBASE up to December 2013. We also searched registers of clinical trials, abstracts of scientific meetings and reference lists of included studies. We searched for randomised controlled trials (RCTs), quasi-RCTs and observational studies that examined women with adult-onset granulosa cell tumours of the ovary (primary and recurrent). For non-randomised studies, we included studies that used multivariate analysis to adjust for baseline characteristics. Two review authors independently abstracted data and assessed risk of bias. Studies were heterogeneous with respect to treatment comparisons, so data were not synthesised in meta-analyses, and methods for assessing heterogeneity were not needed. Risk of bias in included studies was assessed by using the six core items used to assess RCTs and by evaluating four additional criteria specifically addressing risk of bias in non-randomised studies. Five retrospective cohort studies (535 women with a diagnosis of GCT) that used appropriate statistical methods

  20. TGF-β signaling controls FSHR signaling-reduced ovarian granulosa cell apoptosis through the SMAD4/miR-143 axis.

    Science.gov (United States)

    Du, Xing; Zhang, Lifan; Li, Xinyu; Pan, Zengxiang; Liu, Honglin; Li, Qifa

    2016-11-24

    Follicle-stimulating hormone receptor (FSHR) and its intracellular signaling control mammalian follicular development and female infertility. Our previous study showed that FSHR is downregulated during follicular atresia of porcine ovaries. However, its role and regulation in follicular atresia remain unclear. Here, we showed that FSHR knockdown induced porcine granulosa cell (pGC) apoptosis and follicular atresia, and attenuated the levels of intracellular signaling molecules such as PKA, AKT and p-AKT. FSHR was identified as a target of miR-143, a microRNA that was upregulated during porcine follicular atresia. miR-143 enhanced pGC apoptosis by targeting FSHR, and reduced the levels of intracellular signaling molecules. SMAD4, the final molecule in transforming growth factor (TGF)-β signaling, bound to the promoter and induced significant downregulation of miR-143 in vitro and in vivo. Activated TGF-β signaling rescued miR-143-reduced FSHR and intracellular signaling molecules, and miR-143-induced pGC apoptosis. Overall, our findings offer evidence to explain how TGF-β signaling influences and FSHR signaling for regulation of pGC apoptosis and follicular atresia by a specific microRNA, miR-143.

  1. Dioxin-like (PCB 126) and non dioxin-like (PCB 153) action on DNA damage and apoptosis in granulosa cells. Preliminary data

    Energy Technology Data Exchange (ETDEWEB)

    Gregoraszczuk, E.L.; Wojtowicz, A. [Lab. of Physiology and Toxicology of Reproduction, Inst. of Zoology, Jagiellonian Univ., Krakow (Poland); Kapiszewska, M.; Magnowska, Z. [Dept. of General Biochemistry, Jagiellonian Univ., Krakow (Poland)

    2004-09-15

    Introduction Approximately 60-90% of all cancer cases are now generally believed to be due to environmental factors, to which humans are exposed by taking food, and water or inhaling air. These chemicals, including polycyclic aromatic hydrocarbons (PAHs), can be subdivided into different classes. TCDD was classified as group (documented carcinogen in humans) Benzo [a] pyrene and benzo [a] anthracene were included into group 2A (probably carcinogen in humans). PCB is not mentioned in this list. The International Agency for Research on Cancer (IARC) maintains a register of human carcinogens and suspected human carcinogens. The capacity of PCBs to induce DNA strand breaks has been studied to a lesser degree and is quite controversial. DNA strand breaks are the potentially mutagenic lesions that have been proposed as a genotoxicity biomarker for the biomonitoring of environmental pollutants. The mutations induced by these lesions in pollutant-exposed populations are believed to be induced by chemical carcinogens. The formation of PCB adducts has been demonstrated and measured in vitro and in vivo 3. The capacity of PCBs to form DNA adducts depends on their metabolic rate, which is determined by the degree of chlorination. The aim of the presented preliminary data was to evaluate the follicular stage-specific action of PCB 126 and PCB 153 on DNA damage and apoptosis in granulosa cells.

  2. Telomere length is short in PCOS and oral contraceptive does not affect the telomerase activity in granulosa cells of patients with PCOS.

    Science.gov (United States)

    Li, Ying; Deng, Bingbing; Ouyang, Nengyong; Yuan, Ping; Zheng, Lingyan; Wang, Wenjun

    2017-07-01

    Our study aimed to investigate the association of telomerase activity (TA) and telomere length (TL) in granulosa cells (GCs) with IVF outcomes of polycystic ovary syndrome (PCOS) patients, and the effects of oral contraceptive pill (OCP) pretreatment on these two parameters. One hundred sixty-three infertile women were enrolled and divided into a PCOS group (n = 65) and a non-PCOS group (n = 98). The PCOS group was further divided into an OCP pretreatment group (n = 35) and a non-OCP pretreatment group (n = 30), a TA PCOS group and 1.118 in non-PCOS group (P = 0.005). The patients with TL ≥1 accounted for 36.9% in PCOS group and 54.1% in non-PCOS group (P = 0.032). The average duration of infertility for PCOS patients was 5 years in TA PCOS patients. Shorter TL was found in PCOS patients. The TA levels did not change significantly in PCOS patients. PCOS patients with a lower TA level and shorter telomeres had an earlier onset of infertility symptoms. No predictive value was found for TA and TL in terms of embryo quality or IVF outcomes in PCOS patients, and no effect OCP pretreatment was observed on either TA and TL.

  3. Influence of mycotoxin zearalenone and its derivatives (alpha and beta zearalenol on apoptosis and proliferation of cultured granulosa cells from equine ovaries

    Directory of Open Access Journals (Sweden)

    Minoia Paolo

    2006-11-01

    Full Text Available Abstract Background The mycotoxin zearalenone (ZEA and its derivatives, alpha and beta-zearalenol (alpha and beta-ZOL, synthesized by genera Fusarium, often occur as contaminants in cereal grains and animal feeds. The importance of ZEA on reproductive disorders is well known in domestic animals species, particularly in swine and cattle. In the horse, limited data are available to date on the influence of dietary exposure to ZEA on reproductive health and on its in vitro effects on reproductive cells. The aim of this study was to evaluate the effects of ZEA and its derivatives, alpha and beta-ZOL, on granulosa cells (GCs from the ovaries of cycling mares. Methods The cell proliferation was evaluated by using the 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT test after 3 days exposure at different concentrations of ZEA and its derivatives (from 1 × 10-7 to 0.1 microM. The apoptosis induction was evaluated after 1 day exposure, by DNA analysis using flow cytometry. Results An increase in cell proliferation with respect to the control was observed in the presence of ZEA at 1 × 10-3 and 1 × 10-4 microM and apoptosis was induced by all mycotoxins at different concentrations. Conclusion The simultaneous presence of apoptosis and proliferation in GC cultures treated with zearalenones could indicate that these mycotoxins could be effective in inducing follicular atresia. These effects of zearalenones may result from both direct interaction with oestrogen-receptors as well as interaction with the enzymes 3alpha (beta-hydroxysteroid dehydrogenase (HSD, involved in the synthesis and metabolism of endogenous steroid hormones. These cellular disturbances, described for the first time in equine GCs cultured in vitro, could be hypothesized as referred to reproductive failures of unknown ethiology in the mare.

  4. Stability of lutein in wholegrain bakery products naturally high in lutein or fortified with free lutein.

    Science.gov (United States)

    Abdel-Aal, El-Sayed M; Young, J Christopher; Akhtar, Humayoun; Rabalski, Iwona

    2010-09-22

    Lutein is a yellow pigment found in common foods that promotes the health of eyes and skin and is associated with reduced risk of age-related macular degeneration and cataracts. In the present study, selected high-lutein wheat and corn were milled into wholegrain flours by two mills to improve flour uniformity. The high-lutein and lutein-fortified wholegrain flours were processed into breads, cookies, and muffins to study lutein stability during baking and subsequent storage. Lutein and its isomers were separated, identified, and quantified by LC-UV/vis and LC-MS following extraction with water-saturated 1-butanol. Baking resulted in a significant reduction in all-trans-lutein and the formation of cis-lutein and cis-zeaxanthin isomers. Subsequent storage at ambient temperature had a slight impact on the content of all-trans-lutein. Effects of processing were more pronounced in lutein-fortified products, and the degradation rate of lutein was influenced by concentration and baking recipe. Fortified cookies and muffins showed greater lutein reduction compared with bread. Despite the significant reduction in lutein, the fortified bakery products still possessed reasonable amounts per serving that would enhance daily intake and consumption of wholegrain foods.

  5. 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.

  6. Exposure of different cryoprotectants and concentrations on diameter and number of the granulosa cells of bovine preantral follicles Exposição de diferentes crioprotetores e concentrações sobre o diâmetro e número de células da granulosa de folículos pré-antrais bovinos

    Directory of Open Access Journals (Sweden)

    Iraceles Aparecida Laura

    2010-03-01

    Full Text Available This study evaluated the effect of glycerol (GLY, dimethyl sulfoxide (DMSO and propylene glycol (PROH in diameter of follicle, of oocyte and of granulosa cells number of preantral follicles cattle. Ovarian fragments were exposed to cryoprotectants at concentrations of 20 and 40% for 10 min. Then, the tissue was fixed in Carnoy and processed by traditional technique of histology. Slides were stained by hematoxylin and eosin and evaluated under optical microscope equipped with eyes wide with morphometric. Data were analyzed by ANOVA and Tukey test. Regarding diameter of the follicle, no difference was found among the control and the groups exposed to the cryoprotectants. However, in relation to the diameter of the oocyte, 40% concentration in all tested cryoprotectants the primary follicles showed reduction in diameter, including 20% of propylene glycol. For primary follicles, only 40% of glycerol presented difference from control group. For the granulosa cells 40% of DMSO and 20 and 40% of PROH showed significant reduction compared to control group, in primary follicles. In conclusion primary follicles were resistant to cryoprotectants effects. For the primary structures, glycerol, dimethyl sulfoxide and propylene glycol led to reduction of diameter of oocyte and number of granulosa cells but not of follicular diameter.O objetivo, no presente estudo, foi avaliar o efeito do glicerol (GLI, dimetilsulfóxido (DMSO e propilenoglicol (PROH sobre o diâmetro do folículo, do ovócito e do número de células da granulosa de folículos pré-antrais bovinos. Fragmentos ovarianos foram imediatamente fixados em Carnoy (controle ou expostos aos crioprotetores nas concentrações de 20 e 40%, por 10min. Após o período de exposição, o tecido ovariano também foi fixado em Carnoy e processado para inclusão em parafina. As secções (5 de tecido ovariano foram coradas em hematoxilina e eosina e analisadas ao microscópio de luz com ocular em escala microm

  7. The anti-Müllerian hormone (AMH) induces forkhead box L2 (FOXL2) expression in primary culture of human granulosa cells in vitro.

    Science.gov (United States)

    Sacchi, Sandro; Marinaro, Federica; Xella, Susanna; Marsella, Tiziana; Tagliasacchi, Daniela; La Marca, Antonio

    2017-09-01

    Anti-Müllerian hormone (AMH) and forkhead box L2 (FOXL2) are two pivotal genes expressed in human granulosa cells (hGCs) where both genes share similar inhibitory functions on activation and follicular growth in order to preserve the ovarian follicle reserve. Furthermore, AMH and FOXL2 contribute to inhibit steroidogenesis, decreasing or preventing the activation of gonadotrophin-dependent aromatase CYP19A1 cytochrome P450 family 19 subfamily A member 1 (CYP19A1). The purpose of this study is to evaluate the role of AMH in regulating the expression of FOXL2. Primary cultures of hGCs were treated with increasing concentrations of recombinant human AMH (rhAMH; range 10-100 ng/ml) for 3 h. Negative controls were performed using corresponding amounts of AMH vehicle. Total RNA or proteins were purified and quantified by spectrophotometry. FOXL2 and CYP19A1 gene expression, normalized by reference gene ribosomal protein S7 (RpS7), was evaluated by RT-qPCR. Each reaction was repeated in triplicate. Statistical analysis was performed. Extracted proteins were analyzed by immunoblot using anti-FOXL2 and anti-β-actin as primary antibodies. rhAMH treatments tested did not modulate the basal expression of aromatase CYP19A1 gene. rhAMH (50 ng/ml) was able to increase FOXL2 gene expression and its intracellular content. This study demonstrated the existence of an AMH-FOXL2 relationship in hGCs. AMH is capable of increasing both gene and protein expression of FOXL2. Because FOXL2 induces AMH transcription, these ovarian factors could be finely regulated by a positive feedback loop mechanism to preserve the ovarian follicle reserve.

  8. HER2 and GATA4 are new prognostic factors for early-stage ovarian granulosa cell tumor—a long-term follow-up study

    International Nuclear Information System (INIS)

    Färkkilä, Anniina; Andersson, Noora; Bützow, Ralf; Leminen, Arto; Heikinheimo, Markku; Anttonen, Mikko; Unkila-Kallio, Leila

    2014-01-01

    Granulosa cell tumors (GCTs) carry a risk of recurrence also at an early stage, but reliable prognostic factors are lacking. We assessed clinicopathological prognostic factors and the prognostic roles of the human epidermal growth factor receptors (HER 2–4) and the transcription factor GATA4 in GCTs. We conducted a long-term follow-up study of 80 GCT patients with a mean follow-up time of 16.8 years. A tumor-tissue microarray was immunohistochemically stained for HER2–4 and GATA4. Expression of HER2–4 mRNA was studied by means of real time polymerase chain reaction and HER2 gene amplification was analyzed by means of silver in situ hybridization. The results were correlated to clinical data on recurrences and survival. We found that GCTs have an indolent prognosis, with 5-year disease-specific survival (DSS) being 97.5%. Tumor recurrence was detected in 24% of the patients at a median of 7.0 years (range 2.6–18 years) after diagnosis. Tumor stage was not prognostic of disease-free survival (DFS). Of the molecular prognostic factors, high-level expression of HER2, and GATA4, and high nuclear atypia were prognostic of shorter DFS. In multivariate analyses, high-level coexpression of HER2 and GATA4 independently predicted DFS (hazard ratio [HR] 8.75, 95% CI 2.20–39.48, P = 0.002). High-level expression of GATA4 also predicted shorter DSS (HR 3.96, 95% CI 1.45–12.57, P = 0.006). In multivariate analyses, however, tumor stage (II–III) and nuclear atypia were independent prognostic factors of DSS. In conclusion HER2 and GATA4 are new molecular prognostic markers of GCT recurrence, which could be utilized to optimize the management and follow-up of patients with early-stage GCTs

  9. Evidence of a local negative role for cocaine and amphetamine regulated transcript (CART, inhibins and low molecular weight insulin like growth factor binding proteins in regulation of granulosa cell estradiol production during follicular waves in cattle

    Directory of Open Access Journals (Sweden)

    Ireland James J

    2006-04-01

    Full Text Available Abstract The ability of ovarian follicles to produce large amounts of estradiol is a hallmark of follicle health status. Estradiol producing capacity is lost in ovarian follicles before morphological signs of atresia. A prominent wave like pattern of growth of antral follicles is characteristic of monotocous species such as cattle, horses and humans. While our knowledge of the role of pituitary gonadotropins in support of antral follicle growth and development is well established, the intrinsic factors that suppress estradiol production and may help promote atresia during follicular waves are not well understood. Numerous growth factors and cytokines have been reported to suppress granulosa cell estradiol production in vitro, but the association of expression of many such factors in vivo with follicle health status and their physiological significance are not clear. The purpose of this review is to discuss the in vivo and in vitro evidence supporting a local physiological role for cocaine and amphetamine regulated transcript, inhibins and low molecular weight insulin like growth factor binding proteins in negative regulation of granulosa cell estradiol production, with emphasis on evidence from the bovine model system.

  10. 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.

  11. Dopamine in human follicular fluid is associated with cellular uptake and metabolism-dependent generation of reactive oxygen species in granulosa cells: implications for physiology and pathology.

    Science.gov (United States)

    Saller, S; Kunz, L; Berg, D; Berg, U; Lara, H; Urra, J; Hecht, S; Pavlik, R; Thaler, C J; Mayerhofer, A

    2014-03-01

    Is the neurotransmitter dopamine (DA) in the human ovary involved in the generation of reactive oxygen species (ROS)? Human ovarian follicular fluid contains DA, which causes the generation of ROS in cultured human granulosa cells (GCs), and alterations of DA levels in follicular fluid and DA uptake/metabolism in GCs in patients with polycystic ovary syndrome (PCOS) are linked to increased levels of ROS. DA is an important neurotransmitter in the brain, and the metabolism of DA results in the generation of ROS. DA was detected in human ovarian homogenates, but whether it is present in follicular fluid and plays a role in the follicle is not known. We used human follicular fluid from patients undergoing in vitro fertilization (IVF), GCs from patients with or without PCOS and also employed mathematical modeling to investigate the presence of DA and its effects on ROS. DA in follicular fluid and GCs was determined by enzyme-linked immunosorbent assay. GC viability, apoptosis and generation of ROS were monitored in GCs upon addition of DA. Inhibitors of DA uptake and metabolism, an antioxidant and DA receptor agonists, were used to study cellular uptake and the mechanism of DA-induced ROS generation. Human GCs were examined for the presence and abundance of transcripts of the DA transporter (DAT; SLC6A3), the DA-metabolizing enzymes monoamine oxidases A/B (MAO-A/B) and catechol-O-methyltransferase and the vesicular monoamine transporter. A computational model was developed to describe and predict DA-induced ROS generation in human GCs. We found DA in follicular fluid of ovulatory follicles of the human ovary and in GCs. DAT and MAO-A/B, which are expressed by GCs, are prerequisites for a DA receptor-independent generation of ROS in GCs. Blockers of DAT and MAO-A/B, as well as an antioxidant, prevented the generation of ROS (P human follicular compartment, functions of DA could only be studied in IVF-derived GCs, which can be viewed as a cellular model for the

  12. Androgen receptor-mediated non-genomic effects of vinclozolin on porcine ovarian follicles and isolated granulosa cells: Vinclozolin and non-genomic effects in porcine ovarian follicles.

    Science.gov (United States)

    Wartalski, Kamil; Knet-Seweryn, Malgorzata; Hoja-Lukowicz, Dorota; Tabarowski, Zbigniew; Duda, Malgorzata

    2016-05-01

    The present study investigated the influence of the androgen receptor (AR) agonists testosterone (T) and dihydrotestosterone (DHT), and vinclozolin (Vnz), a fungicide with antiandrogenic activity, on non-genomic signal transduction within ovarian follicles. Porcine granulosa cells (GCs) isolated from mature follicles were cultured for 48h. For the last 24h of culture, they were exposed to T (10(-7)M), DHT (10(-7)M), Vnz (1.4×10(-5)M), T and Vnz (T+Vnz), or DHT and Vnz (DHT+Vnz) at the same concentrations. To better imitate in vivo conditions, whole follicles (4-6mm in diameter) were incubated (24h) in an organ culture system with the same factors. Expression of AR mRNA and protein was determined by real-time PCR and western blot analyses. To demonstrate AR localization in cultured GCs and whole follicles, immunocytochemistry and immunohistochemistry were performed, respectively. To elucidate the possible non-genomic action of Vnz in GCs, protein expression and the activity of ERK1/2 and Akt kinases were determined by western blot and ELISA analyses. The immunocytochemistry and immunohistochemistry results showed that exposure of GCs and follicles to Vnz resulted in cytoplasmic and perinuclear AR localization. Real-time PCR and western blot analysis showed that AR mRNA and protein expression increased (P≤0.001) in GC cultures after combined treatment with an androgen and Vnz. In whole follicles, such treatment also increased AR mRNA with a decrease in the respective protein expression (P≤0.001). Moreover, addition of T or DHT with Vnz increased the activity of ERK1/2 and Akt kinases in cultured GCs (P≤0.001). The results suggest a novel mechanism for Vnz action in porcine ovarian follicles on both AR mRNA and protein levels. Thus, this environmental antiandrogen activates non-genomic signaling pathways, as indicated by the increased activity of both investigated kinases observed within minutes of Vnz addition. Given the widespread presence of Vnz in the

  13. 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

  14. Influence of Estradiol-17beta on Progesterone and Estrogen Receptor mRNA Expression in Porcine Follicular Granulosa Cells during Short-Term, In Vitro Real-Time Cell Proliferation

    Directory of Open Access Journals (Sweden)

    Sylwia Ciesiółka

    2016-01-01

    Full Text Available Progesterone (P4 and estradiol (E2 play a significant role in mammalian reproduction. Our study demonstrated that separated porcine cumulus cells (CCs and/or granulosa cells (GCs might proliferate in vitro during short-term, real-time primary culture. The GCs were analyzed according to gene expression of the progesterone receptor (nuclear form (pgr, progesterone receptor membrane component 1 (pgrmc1, and estrogen-related receptor beta 3 (esrrb3 in relation to two housekeeping genes: actb and pbgd. GCs were cultivated in medium with the E2. Both pgr/actb and pgr/pbgd revealed higher expression between 24 and 168 h of IVC of prolonged E2 treatment and at 48 h of IVC after acute E2 administration. The pgrmc1/actb and pgrmc1/pbgd displayed increased expression after prolonged E2 treatment between 24 and 120 h of IVC. The highest level of esrrb3/actb at 120 and 144 h, as well as esrrb3/pbgd at 120 h, in untreated controls as compared to the hormone-stimulated group, was observed. We suggest that E2 significantly influences the upregulation of pgr, pgrmc1, and esrrb3 expression in porcine GCs during real-time cell proliferation. Since esrrb3 expression is stimulated by E2 in both an acute and prolonged manner, estradiol may be recognized as a potential estrogen receptor agonist in GCs.

  15. Morphometry and granulosa cells number of preantral ovarian follicles from bovine (“Bos indicus” preserved at 4ºC in saline solution for different periods of time Morfometria e número de células da granulosa de folículos ovarianos pré-antrais de bovino ("Bos indicus" preservados a 4ºC em solução salina por diferentes períodos de tempo

    Directory of Open Access Journals (Sweden)

    Hélder Silva Luna

    2008-04-01

    Full Text Available The present study had the objective to evaluate the effect of cooling at 4 ºC, at different periods of time, in the morphometry and number of granulosa cells of preantral ovarian follicles of bovines. Ovaries were gotten from slaughterhouse, divided in 4 parts and distributed in 4 groups: control (without cooling and groups cooled for 12, 24 and 48 h in saline solution (NaCl 0.9% at 4 ºC. After treatment, samples were fixed in Carnoy and forwarded to classic histology techniques. The follicles were analyzed with optic and ocular microscope with micrometric scale. Total of 340 follicles were analyzed, which 242 were primordial and 98 primary. Results indicated that the primordial and primary follicles when cooled in saline solution at 4 ºC did not present variation in oocyte and diameter and amount of cells of the granulosa when comparing the control group to 12, 24 and 48 h groups (P > 0.05. In conclusion, the temperature at 4 ºC does not influence the morphometry of the follicles and the amount of the granulosa cells.O presente trabalho teve como objetivo avaliar o efeito do resfriamento a 4 ºC, em diferentes períodos de tempo, na morfometria e número de células da granulosa de folículos pré-antrais de ovário bovino. Os ovários foram obtidos em abatedouro, divididos em 4 partes e distribuídos em 4 grupos: controle (sem resfriamento e grupos resfriados por 12, 24 e 48 h em solução salina (NaCl a 0,9% a 4 ºC. Após tratamento, as amostras foram fixadas em Carnoy e encaminhadas para rotina histológica clássica. Para análise folicular, utilizou-se microscópio óptico e ocular com escala micrométrica. Foram analisados um total de 340 folículos, sendo 242 primordiais e 98 primários. Os resultados obtidos indicam que os folículos primordiais e primários, quando resfriados em solução salina a 4 ºC, não apresentam variação em sua morfometria e quantidade de células da granulosa, quando comparado o grupo controle com

  16. The polymorphic insertion of the luteinizing hormone receptor "insLQ" show a negative association to LHR gene expression and to the follicular fluid hormonal profile in human small antral follicles

    DEFF Research Database (Denmark)

    Borgbo, T; Chrudimska, J; Macek, M

    2018-01-01

    (AMHR2) and LHCGR, respectively, were observed for insLQ/insLQ compared to -/insLQ and the -/- genotypes. Moreover, LHCGR and CYP19a1 together with oestradiol and inhibin-B were significantly increased in -/insLQ compared to the -/- genotype. The homozygous insLQ genotype showed strong significant......The luteinizing hormone receptor (LHCGR) has a little studied polymorphic 6 bp insertion (rs4539842/insLQ). This study has evaluated the insLQ polymorphism in relation to potential associations with hormonal characteristics of human small antral follicles (hSAFs). In total, 310 hSAFs were collected...... from 86 women undergoing fertility preservation. Analysis included hormonal profile of 297 follicular fluid (FF) samples and 148 corresponding granulosa cells samples were evaluated by qPCR for selected genes. Significantly reduced and non-detectable mRNA levels of anti-Müllerian hormone receptor II...

  17. 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...... was significantly increased, whereas AMH gene expression was significantly reduced for the G/G genotype. In follicles >6 mm, estradiol and CYP19A1 gene expression levels were significantly higher for the G/G genotype. In conclusion, significant changes were observed between the FSHR 307/680 polymorphisms in human......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...

  18. Bioavailability and biodistribution of nanodelivered lutein

    Science.gov (United States)

    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 ...

  19. Relation among serum and tissue concentrations of lutein and zeaxanthin and macular pigment density.

    Science.gov (United States)

    Johnson, E J; Hammond, B R; Yeum, K J; Qin, J; Wang, X D; Castaneda, C; Snodderly, D M; Russell, R M

    2000-06-01

    Lutein and zeaxanthin are the only carotenoids in the macular region of the retina (referred to as macular pigment [MP]). Foods that are rich in lutein and zeaxanthin can increase MP density. Response to dietary lutein and zeaxanthin in other tissues has not been studied. The objective of this study was to examine tissue responses to dietary lutein and zeaxanthin and relations among tissues in lutein and zeaxanthin concentrations. Seven subjects consumed spinach and corn, which contain lutein and zeaxanthin, with their daily diets for 15 wk. At 0, 4, 8, and 15 wk and 2 mo after the study, serum, buccal mucosa cells, and adipose tissue were analyzed for carotenoids, and MP density was measured. Serum and buccal cell concentrations of lutein increased significantly from baseline during dietary modification. Serum zeaxanthin concentrations were greater than at baseline only at 4 wk, whereas buccal cell and adipose tissue concentrations of zeaxanthin did not change. Adipose tissue lutein concentrations peaked at 8 wk. Changes in adipose tissue lutein concentration were inversely related to the changes in MP density, suggesting an interaction between adipose tissue and retina in lutein metabolism. To investigate the possibility of tissue interactions, we examined cross-sectional relations among serum, tissue, and dietary lutein concentrations, anthropometric measures, and MP density in healthy adults. Significant negative correlations were found between adipose tissue lutein concentrations and MP for women, but a significant positive relation was found for men. Sex differences in lutein metabolism may be an important factor in tissue interactions and in determining MP density.

  20. 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.

  1. Evidence for gating roles of protein kinase A and protein kinase C in estradiol-induced luteinizing hormone receptor (lhcgr) expression in zebrafish ovarian follicle cells.

    Science.gov (United States)

    Liu, Ka-Cheuk; Ge, Wei

    2013-01-01

    Estradiol (E2) stimulates luteinizing hormone receptor (lhcgr) expression in zebrafish follicle cells via nuclear estrogen receptors (nERs) that are likely expressed on the membrane, and lhcgr responds to E2 in a biphasic manner during 24-h treatment. These observations raise an interesting question on the signaling mechanism underlying E2 regulation, in particular the biphasic response of lhcgr expression. In the present study, we demonstrated that E2 regulation of lhcgr was significantly influenced by the activity of cAMP-PKA pathway. Activation of cAMP-PKA pathway by forskolin or db-cAMP suppressed E2-stimulated lhcgr expression in short-term (3 h) but enhanced its effect in long-term (24 h), suggesting differential roles of PKA at these two phases of lhcgr response. PKA inhibitor H89 showed reversed effects. In contrast, PKC pathway had consistent permissive effect on E2-induced lhcgr expression as evidenced by strong inhibition of E2 effect by PKC inhibitors GF109203X and Ro-31-8220 at both 3 and 24 h. One of the mechanisms by which PKA and PKC gated E2 effect might be through regulating nERs, particularly esr2a. Despite the strong influence of PKA and PKC, our data did not suggest direct mediating roles for these two pathways in E2 stimulation of lhcgr expression; yet they likely play critical gating roles in E2 signal transduction. As a follow-up study to our previous report on E2 regulation of gonadotropin receptors in the zebrafish ovary, the present study provides further evidence for the involvement of classical intracellular signal transduction pathways in E2 stimulation of lhcgr expression in the follicle cells.

  2. Lutein and atherosclerosis: Belfast versus Toulouse revisited.

    Science.gov (United States)

    Howard, A N; Thurnham, D I

    2017-01-01

    In 1995 we reported that mean plasma lutein concentrations in salaried men and women from Toulouse in Southern France were double those in subjects recruited from general practitioner lists in Belfast, Northern Ireland. At the time incidence of coronary heart disease (CHD) in Southern France was among the lowest in Europe and was much higher in Northern Ireland. Plasma lutein is a biomarker of vegetable and fruit intake and evidence suggests that high concentrations are generally associated with better cardiometabolic health. At the time we speculated like others that role of the carotenoids may well have been to prevent oxidation of lipid in the lipoproteins and so reduce the uptake of oxidised lipid by macrophages and its deposition within the intimal layers of the major arteries as plaque. It is now widely accepted that CHD is an inflammatory disease and that macrophages within plaque together with tissue damage contribute to this inflammation. Stimulated macrophages release cytokines to activate the immune system both locally and systemically. Precursor complement proteins in the blood are activated to assist immune cells in phagocytosis and cell repair. Individuals with a history of arteriosclerosis display significantly higher concentrations of complement factors C3 and C3a than subjects without such a history. Metabolism of C3 via the alternate complement pathway can give rise to the membrane attack complex (MAC) which creates a hole or pore in pathogens or host cells, killing the cell. Recent studies in patients with early age related macular disease (AMD) who also exhibit similar elevated concentrations of complement proteins in their blood, showed supplementation with lutein progressively decreased the amount of the MAC and other complement factors in the blood. Lutein was used in the supplementation experiments because it is an important constituent of macular pigment. Thus the healthier cardiometabolic features displayed by the people in Toulouse may

  3. Lutein bioavailability is higher from lutein-enriched eggs than from supplements and spinach in men.

    Science.gov (United States)

    Chung, Hae-Yun; Rasmussen, Helen M; Johnson, Elizabeth J

    2004-08-01

    Lutein may be protective against diseases such as age-related macular degeneration (ARMD). At present, data regarding bioavailability of lutein from various sources are insufficient. Healthy men (n = 10) participated in an intervention study with a crossover design. After a 2-wk washout period during which they consumed a low-carotenoid diet, the men were administered 1 of 4 lutein doses (lutein supplement, lutein ester supplement, spinach, and lutein-enriched egg) for 9 d. All lutein doses provided 6 mg lutein except for the lutein ester dose, which provided 5.5 mg lutein equivalents. Serum samples were collected from fasting subjects on d -14, 1 (baseline), 2, 3, and 10 and analyzed for changes in lutein concentration. Triacylglycerol-rich lipoproteins (TRL) were separated from postprandial blood samples (0-24 h) after the first lutein dose and analyzed for lutein concentration. Subjects completed all 4 treatments of the study in random order. Results from repeated-measures 1-way ANOVA showed that the baseline and dose-adjusted lutein response in serum was significantly higher after egg consumption than after lutein, lutein ester, and spinach consumption on d 10. There was no significant difference in TRL response. In conclusion, the lutein bioavailability from egg is higher than that from other sources such as lutein, lutein ester supplements, and spinach. The lutein bioavailability from lutein, lutein ester supplements, and spinach did not differ. This finding may have implications for dietary recommendations that may decrease the risk of certain diseases, e.g., ARMD.

  4. Serum lutein response is greater from free lutein than from esterified lutein during 4 weeks of supplementation in healthy adults.

    Science.gov (United States)

    Norkus, Edward P; Norkus, Katherine L; Dharmarajan, T S; Schierle, Joseph; Schalch, Wolfgang

    2010-12-01

    Current data suggest great variability in serum response following lutein ingestion from various sources. To compare the relative serum response during supplementation with free lutein (fL) and lutein esters (Le). 72 volunteers (23-52 years; body mass index [BMI] >20 and lutein lutein or 27 mg of lutein ester (equivalent to 13.5 mg free lutein), respectively. Fasting blood was obtained at baseline and after 7, 14, 21, and 28 days of supplementation. Supplements were consumed with standard portions of dry, ready-to-eat cereal and 2% cow's milk. Absolute changes in serum lutein, per mg daily dose, were significantly greater in fL vs. Le after 21 days (p  =  0.0012) and remained so after 28 days (p  =  0.0011) of supplementation. Serum lutein Area Under the Curve [AUC((day 0-28))] response was 17% greater for fL vs. Le (p  =  0.0187). Regression models were used and determined that (1) baseline serum lutein levels and (2) the form of lutein ingested (fL > Le) influence the serum lutein response during supplementation, while subject age, gender, BMI, and serum lipids do not affect serum response. These results suggest that the relative serum lutein response will be significantly greater from supplements containing free lutein than from supplements containing lutein esters. These findings should be useful for future clinical trials exploring the effectiveness of lutein supplementation in the prevention of or protection against age-related macular degeneration and/or cataracts.

  5. 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

    changes of circulating estradiol (E2) and progesterone (P4) levels after administration of hCG. In vitro, when MGCs and COCs were treated with U0126, a specific inhibitor of MAPK3/1 activation, gonadotropin- or 8-Br-cAMP-induced P4 production, as well as expression of Star and Cyp11a1 mRNA, were...

  6. Lutein facilitates physiological revascularization in a mouse model of retinopathy of prematurity.

    Science.gov (United States)

    Fu, Zhongjie; Meng, Steven S; Burnim, Samuel B; Smith, Lois Eh; Lo, Amy Cy

    2017-07-01

    Retinopathy of prematurity is one of the leading causes of childhood blindness worldwide, with vessel growth cessation and vessel loss in phase I followed by neovascularization in phase II. Ischaemia contributes to its pathogenesis, and lutein protects against ischaemia-induced retinal damages. We aimed to investigate the effects of lutein on a murine model of oxygen-induced retinopathy. Mouse pups were exposed to 75% oxygen for 5 days and returned to room air for another 5 days. Vascular obliteration, neovascularization and blood vessel leakage were examined. Immunohistochemistry for glial cells and microglia were performed. Compared with vehicle controls, mouse pups receiving lutein treatment displayed smaller central vaso-obliterated area and reduced blood vessel leakage. No significant difference in neovascular area was found between lutein and vehicle controls. Lutein promoted endothelial tip cell formation and maintained the astrocytic template in the avascular area in oxygen-induced retinopathy. No significant changes in Müller cell gliosis and microglial activation in the central avascular area were found in lutein-treated pups. Our observations indicated that lutein significantly promoted normal retinal vascular regrowth in the central avascular area, possibly through promoting endothelial tip cell formation and preserving astrocytic template. Our results indicated that lutein might be considered as a supplement for the treatment of proliferative retinopathy of prematurity because of its role in facilitating the revascularization of normal vasculature. © 2016 Royal Australian and New Zealand College of Ophthalmologists.

  7. Serum lutein concentrations in healthy term infants fed human milk or infant formula with lutein

    OpenAIRE

    Bettler, Jodi; Zimmer, J. Paul; Neuringer, Martha; DeRusso, Patricia A.

    2009-01-01

    Background Lutein is a carotenoid that may play a role in eye health. Human milk typically contains higher concentrations of lutein than infant formula. Preliminary data suggest there are differences in serum lutein concentrations between breastfed and formula-fed infants. Aim of the study To measure the serum lutein concentrations among infants fed human milk or formulas with and without added lutein. Methods A prospective, double-masked trial was conducted in healthy term formula-fed infant...

  8. Caco-2 accumulation of lutein is greater from human milk than from infant formula despite similar bioaccessibility.

    Science.gov (United States)

    Lipkie, Tristan E; Banavara, Dattatreya; Shah, Bhavini; Morrow, Ardythe L; McMahon, Robert J; Jouni, Zeina E; Ferruzzi, Mario G

    2014-10-01

    Clinical evidence suggests that the bioavailability of lutein is lower from infant formula than from human milk. The purpose of this study was to assess characteristics of human milk and lutein-fortified infant formula that may impact carotenoid delivery. Carotenoid bioaccessibility and intestinal absorption were modeled by in vitro digestion coupled with Caco-2 human intestinal cell culture. Twelve human milk samples were assessed from 1-6 months postpartum, and 10 lutein-fortified infant formula samples from three lutein sources in both ready-to-use and reconstituted powder forms. The relative bioaccessibility of lutein was not different (p > 0.05) between human milk (29 ± 2%) and infant formula (36 ± 4%). However, lutein delivery was 4.5 times greater from human milk than infant formula when including Caco-2 accumulation efficiency. Caco-2 accumulation of lutein was increasingly efficient with decreasing concentration of lutein from milk. Carotenoid bioaccessibility and Caco-2 accumulation were not affected by lactation stage, total lipid content, lutein source, or form of infant formula (powder vs. liquid). These data suggest that the bioavailability of carotenoids is greater from human milk than infant formula primarily due to intestinal absorptive processes, and that absorption of lutein is potentiated by factors from human milk especially at low lutein concentration. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Overexpression of miR-21 in stem cells improves ovarian structure and function in rats with chemotherapy-induced ovarian damage by targeting PDCD4 and PTEN to inhibit granulosa cell apoptosis.

    Science.gov (United States)

    Fu, Xiafei; He, Yuanli; Wang, Xuefeng; Peng, Dongxian; Chen, Xiaoying; Li, Xinran; Wang, Qing

    2017-08-14

    Chemotherapy-induced premature ovarian failure (POF) is a severe complication affecting tumor patients at a childbearing age. Mesenchymal stem cells (MSCs) can partially restore the ovarian structure and function damaged by chemotherapy. miR-21 is a microRNA that can regulate cell apoptosis. This study discusses the repair effect and mechanism of MSCs overexpressing miR-21 on chemotherapy-induced POF. Rat MSCs and granulosa cells (GCs) were isolated in vitro. MSCs were transfected with miR-21 lentiviral vector (LV-miR-21) to obtain MSCs stably expressing miR-21 (miR-21-MSCs). The microenvironment of an ovary receiving chemotherapy was mimicked by adding phosphamide mustard (PM) into the cellular culture medium. The apoptosis rate and the mRNA and protein expression of target genes PTEN and PDCD4 were detected in MSCs. Apoptosis was induced by adding PM into the culture medium for GCs, which were cocultured with miR-21-MSCs. The apoptosis rate and the mRNA and protein expression of PTEN and PDCD4 were detected. The chemotherapy-induced POF model was built into rats by intraperitoneal cyclophosphamide injection. miR-21-MSCs were transplanted into the bilateral ovary. The rats were sacrificed at 15, 30, 45, and 60 days after the last injection. The ovarian weights, follicle count, estrous cycle, and sex hormone levels (estradiol (E2) and follicle-stimulating hormone (FSH)) were detected. Apoptosis of GCs was determined by TUNEL assay. The miR-21 and mRNA and protein expression of PTEN and PDCD4 were determined. The apoptosis decreased in MSCs transfected with miR-21. The mRNA and protein expression of target genes PTEN and PDCD4 was downregulated. GCs cocultured with miR-21-MSCs showed a decreased apoptosis, an upregulation of miR-21, and a downregulation of PTEN and PDCD4. Following the injection of miR-21-MSCs, the ovarian weight and follicle counts increased; E 2 levels increased while FSH levels decreased, with less severe apoptosis of GCs. The miR-21 expression

  10. Influência de diferentes concentrações de etileno glicol nO número de células da granulosa e morfometria de folículos pré-antrais inclusos em tecido ovariano de Bos taurus indicus, Linnaeus, 1758 Influence of different concentrations of the glycol in the number of granulosa cells and morphometry of the preantral ovarian follicles of Bos taurus indicus, Linnaeus, 1758

    Directory of Open Access Journals (Sweden)

    Hélder Silva e Luna

    2010-12-01

    Full Text Available A criopreservação de folículos ovarianos pré-antrais pode ajudar na conservação de muitas espécies domésticas e selvagens. Neste trabalho, objetivou-se verificar o efeito do etileno glicol, em diferentes concentrações, na morfometria e número de células da granulosa de folículos pré-antrais inclusos em tecido ovariano bovino. O teste de toxicidade foi realizado com fragmentos ovarianos expostos ao etileno glicol em concentrações de 10, 20 ou 40%. O tecido foi analisado por técnica histológica clássica. Os folículos primordiais expostos à concentração de 40% apresentaram redução do diâmetro folicular e ovocitário quando comparados ao grupo controle (sem exposição, 10% e 20% (P0,05. Esses resultados sugerem que folículos primários são mais resistentes aos efeitos do etileno glicol quando comparados aos primordiais.The cryopreservation of the ovarian preantral follicles could help the conservation of several domestic and wild animal species. The objective of this study was to verify the effect of different concentrations of ethylene glycol on the morphometry and number of granulosa cells of the preantral ovarian follicles. A toxicity test was conducted with strips of ovarian cortex using ethylene glycol (10, 20 or 40%. Tissue analysis was done using classical histology techniques. The primordial follicles expose at a concentration of 40% showed reduction of the follicular diameter and oocyte when compared to the control (non exposing, 10 and 20% groups (P0.05. These results suggest that primary follicles are more resistant to the effect of ethylene glycol when compared to primordial ones.

  11. Characterization of milk proteins-lutein complexes and the impact on lutein chemical stability.

    Science.gov (United States)

    Yi, Jiang; Fan, Yuting; Yokoyama, Wallace; Zhang, Yuzhu; Zhao, Liqing

    2016-06-01

    In this study, the interaction of WPI (whey protein isolate) and SC (sodium caseinate) with hydrophobic lutein was investigated through UV-vis spectroscopy and circular dichroism (CD) as well as fluorescence. The effects on lutein's chemical stability were also examined. The decrease of turbidity of lutein suggested that lutein's aqueous solubility was improved after binding with milk proteins. CD analysis indicated lutein had little impact on the secondary structures of both proteins. Different preparation methods have significant impacts on the binding constant. Fluorescence results indicated that WPI and SC interact with lutein by hydrophobic contacts. Milk proteins have protective effects on lutein against oxidation and decomposition, and SC showed better capability in protecting lutein from oxidation than WPI during 16 days storage. The lutein's chemical stability was increased with increasing of proteins concentration. The results indicated that milk proteins may act as effective carriers for lipophilic nutraceuticals. Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. Effect of bovine pellucid zone 3 monoclonal antibodies on B cell lymphoma 2 expressions of granulosa cell and mice (Mus musculus follicle diameter

    Directory of Open Access Journals (Sweden)

    Heti Ira Ayue

    2017-01-01

    Full Text Available Objective: To evaluate the effects of pellucid zone 3 monoclonal antibodies against B-cell lymphoma 2 (BCL-2 expression and mice follicle diameter at various time periods.Methods: The animal model of this study was 36 Balb/c mice (Mus musculus. A true experimental design was used with a post-test only control group approach. BCL-2 expression was observed using immunohistochemistry, while the follicle diameter was observed by haematoxylin-eosin staining. The data was analyzed using nested ANOVA to compare the results of the mean expression of BCL-2 on the 5th and 20th day of observation in the pre-antral and antral follicle between the control and treatment groups.Results: No significant differences were found in BCL-2 gene expression. There were also no significant differences in BCL-2 expression on the 10th day of pre-antral follicle analysis. Moreover, there were no significant differences between the mean follicle diameter on the 5th, 10th, and 20th day of pre-antral and antral follicle development between the control and treatment groups. The addition of bovine pellucid zone 3 (bZP3 monoclonal antibodies on the 5th and 20th day of observation did not decrease the expression of BCL-2 gene in the pre-antral and antral follicle of mice. Administering bZP3 monoclonal antibodies on the 10th day of observation did not affect BCL-2 expression in the pre-antral follicle but did decrease BCL-2 expression in the antral follicle. Supplying bZP3 monoclonal antibodies on the 5th, 10th and 20th day did not affect the diameter of pre-antral and antral follicles of the mice.Conclusion: The monoclonal antibodies bovine zona pelusida 3 has the potential to be developed as a safe immunocontraception preparation.

  13. Prohibitin (PHB) inhibits apoptosis in rat granulosa cells (GCs) through the extracellular signal-regulated kinase 1/2 (ERK1/2) and the Bcl family of proteins.

    Science.gov (United States)

    Chowdhury, Indrajit; Thompson, Winston E; Welch, Crystal; Thomas, Kelwyn; Matthews, Roland

    2013-12-01

    Mammalian ovarian follicular development is tightly regulated by crosstalk between cell death and survival signals, which include both endocrine and intra-ovarian regulators. Whether the follicle ultimately ovulates or undergoes atresia is dependent on the expression and actions of factors promoting follicular cell proliferation, differentiation or apoptosis. Prohibitin (PHB) is a highly conserved, ubiquitous protein that is abundantly expressed in granulosa cells (GCs) and associated with GC differentiation and apoptosis. The current study was designed to characterize the regulation of anti-apoptotic and pro-apoptotic factors in undifferentiated rat GCs (gonadotropin independent phase) governed by PHB. Microarray technology was initially employed to identify potential apoptosis-related genes, whose expression levels within GCs were altered by either staurosporine (STS) alone or STS in presence of ectopically over-expressed PHB. Next, immunoblot studies were performed to examine the expression patterns of selective Bcl-2 family members identified by the microarray analysis, which are commonly regulated in the intrinsic-apoptotic pathway. These studies were designed to measure protein levels of Bcl2 family in relation to expression of the acidic isoform (phosphorylated) PHB and the components of MEK-Erk1/2 pathway. These studies indicated that over-expression of PHB in undifferentiated GCs inhibit apoptosis which concomitantly results in an increased level of the anti-apoptotic proteins Bcl2 and Bclxl, reduced release of cytochrome c from mitochondria and inhibition of caspase-3 activity. In contrast, silencing of PHB expression resulted in change of mitochondrial morphology from the regular reticular network to a fragmented form, which enhanced sensitization of these GCs to the induction of apoptosis. Collectively, these studies have provided new insights on the PHB-mediated anti-apoptotic mechanism, which occurs in undifferentiated GCs through a PHB → Mek-Erk1

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

    Directory of Open Access Journals (Sweden)

    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.

  15. Paraqueratosis granulosa de la axila.

    Directory of Open Access Journals (Sweden)

    Gerzaín Rodríguez

    2002-12-01

    Full Text Available La paraqueratosis granulosa de la axila es un transtorno adquirido de la cornificación, en el cual la capa córnea es muy gruesa y conserva los gránulos de profilagrina (queratohialina. Se han publicado 32 casos desde su descripción en 1991, de Estados Unidos, Europa y Australia. No conocemos informes de la entidad en Latinoamérica. Estudiamos tres mujeres obesas que presentaron pápulas y placas axilares, costrosas, hiperqueratósicas e hiperpigmentadas, que se diagnosticaron clínicamente como pénfigo familiar benigno o tinea nigra. Las biopsias mostraron enorme capa córnea paraqueratósica con aspecto basófilo dado por la presencia de gránulos de queratohialina, identificados con la hematoxilina-eosina y con microscopía electrónica del tejido obtenido del bloque de parafina. La inmunohistoquímica para S-100 no demostró células de Langerhans en las lesiones. La coloración de PAS no demostró hongos. Los infundíbulos presentaron voluminosos tapones córneos con el mismo cambio. No se vió mayor inflamación dérmica. Estos hallazgos sugieren que la entidad es una dermatitis de contacto irritativo. Las historias clínicas y la revisión de la literatura indican que el uso de desodorantes, la obesidad y la humedad son los factores desencadenantes. El diagnóstico debe ser realizado sin dificultad por el patólogo o el dermatopatólogo, ya que por ser una entidad relativamente nueva no es fácil reconocerla clínicamente.

  16. Enhancing lutein productivity of an indigenous microalga Scenedesmus obliquus FSP-3 using light-related strategies.

    Science.gov (United States)

    Ho, Shih-Hsin; Chan, Ming-Chang; Liu, Chen-Chun; Chen, Chun-Yen; Lee, Wen-Lung; Lee, Duu-Jong; Chang, Jo-Shu

    2014-01-01

    Lutein, one of the main photosynthetic pigments, is a promising natural product with both nutritional and pharmaceutical applications. In this study, light-related strategies were applied to enhance the cell growth and lutein production of a lutein-rich microalga Scenedesmus obliquus FSP-3. The results demonstrate that using white LED resulted in better lutein production efficiency when compared to the other three monochromatic LEDs (red, blue, and green). The lutein productivity of S. obliquus FSP-3 was further improved by adjusting the type of light source and light intensity. The optimal lutein productivity of 4.08 mg/L/d was obtained when using a TL5 fluorescent lamp at a light intensity of 300 μmol/m(2)/s, and this performance is better than that reported in most related studies. Moreover, the time-course profile of lutein accumulation in the microalga shows that the maximal lutein content and productivity were obtained at the onset of nitrogen depletion. Copyright © 2013 Elsevier Ltd. All rights reserved.

  17. Serum lutein concentrations in healthy term infants fed human milk or infant formula with lutein.

    Science.gov (United States)

    Bettler, Jodi; Zimmer, J Paul; Neuringer, Martha; DeRusso, Patricia A

    2010-02-01

    Lutein is a carotenoid that may play a role in eye health. Human milk typically contains higher concentrations of lutein than infant formula. Preliminary data suggest there are differences in serum lutein concentrations between breastfed and formula-fed infants. To measure the serum lutein concentrations among infants fed human milk or formulas with and without added lutein. A prospective, double-masked trial was conducted in healthy term formula-fed infants (n = 26) randomized between 9 and 16 days of age to study formulas containing 20 (unfortified), 45, 120, and 225 mcg/l of lutein. A breastfed reference group was studied (n = 14) and milk samples were collected from their mothers. Primary outcome was serum lutein concentration at week 12. Geometric mean lutein concentration of human milk was 21.1 mcg/l (95% CI 14.9-30.0). At week 12, the human milk group had a sixfold higher geometric mean serum lutein (69.3 mcg/l; 95% CI 40.3-119) than the unfortified formula group (11.3 mcg/l; 95% CI 8.1-15.8). Mean serum lutein increased from baseline in each formula group except the unfortified group. Linear regression equation indicated breastfed infants had a greater increase in serum lutein (slope 3.7; P milk lutein than formula-fed infants (slope 0.9; P lutein concentrations than infants who consume formula unfortified with lutein. These data suggest approximately 4 times more lutein is needed in infant formula than in human milk to achieve similar serum lutein concentrations among breastfed and formula fed infants.

  18. MicroRNAs control transcription factor NF-kB (p65) expression in human ovarian cells.

    Science.gov (United States)

    Sirotkin, Alexander V; Alexa, Richard; Kišová, Gabriela; Harrath, Abdel Halim; Alwasel, Saleh; Ovcharenko, Dmitriy; Mlynček, Miloš

    2015-05-01

    MicroRNAs (miRNAs) are known to influence ovarian cell proliferation, apoptosis and hormone release, but it remains unknown whether miRNAs affect ovarian functions via transcription factors. We examined the effect of miRNAs on nuclear factor-κappaB (NF-kB) (p65) expression in human ovarian luteinized granulosa cells. We transfected cultured primary human ovarian luteinized granulosa cells with 80 different constructs encoding human pre-miRNAs and then evaluated NF-kB (p65) expression (percentage of cells containing p65) by immunocytochemistry. We found that 21 of the constructs stimulated NF-kB (p65) expression and 18 of the constructs inhibited NF-kB (p65) expression. This is the first direct demonstration that miRNAs affect NF-kB (p65) expression and the first genome-scale miRNA screen to identify upregulation and downregulation of NF-kB accumulation by miRNAs in the ovary. Novel miRNAs that affect the NF-kB signalling pathway could be useful for the control of NF-kB-dependent reproductive processes and the treatment of NF-kB-dependent reproductive disorders.

  19. Luteinizing hormone in testicular descent

    DEFF Research Database (Denmark)

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

    2007-01-01

    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....... 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...

  20. Lutein concentration in human milk during early lactation and its relationship with dietary lutein intake.

    Science.gov (United States)

    Cena, Hellas; Castellazzi, Anna Maria; Pietri, Amedeo; Roggi, Carla; Turconi, Giovanna

    2009-10-01

    The present study aimed to estimate the lutein concentration in human milk during early lactation and its relationship with dietary lutein intake measured through the administration of a short FFQ. A cross-sectional study in which an FFQ was administered twice: on day 3 (T0) and day 30 (T1) postpartum; meanwhile two breast milk samples were collected. Maternal plasma samples were obtained at T0. The comparison of dietary lutein intakes and likewise lutein concentrations in breast milk at T0 and T1 were analysed with Student's t test. Pearson's correlation coefficient was used to determine the association between dietary lutein intake and lutein concentration in milk and plasma, respectively, as well as the correlation between breast milk and plasma lutein concentrations at T0. Pavia, northern Italy. Twenty-one pregnant women, age range 24-42 years, were recruited during their last trimester on a voluntary basis. Both breast milk and plasma lutein concentrations were significantly correlated with dietary lutein intake (r = 0.86, P = 0.0001 and r = 0.94, P = 0.0001, respectively). There was a clear significant correlation between milk and plasma lutein concentrations (r = 0.87, P = 0.0001). Mature milk lutein concentration, although significantly reduced at T1 (P lutein intake (r = 0.82, P = 0.0001). Even though milk lutein concentration decreased during early lactation, it remained significantly correlated with daily lutein intake. Therefore, while awaiting further research, dietary recommendations advising intake of fresh fruit and vegetables rich in lutein, throughout the whole duration of pregnancy and lactation, are extremely useful.

  1. Lutein

    Science.gov (United States)

    ... disorder in premature infants that can lead to blindness (retinopathy of prematurity). Research shows that giving preterm ... mg of zeaxanthin does not improve speaking or memory in older people. However, other early research suggests ...

  2. 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.

  3. 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.

  4. Cellular transport of lutein is greater from uncooked rather than cooked spinach irrespective of whether it is fresh, frozen, or canned.

    Science.gov (United States)

    O'Sullivan, Laurie; Ryan, Lisa; Aherne, S Aisling; O'Brien, Nora M

    2008-08-01

    Lutein, a carotenoid found in significant levels in spinach, has attracted a great deal of attention owing to its reported function as a shield against the photooxidative effects of blue light. Therefore, the rationale of this study was to examine the effects of various processing and cooking methods on lutein bioavailability from spinach (Spinacia oleracea) using an in vitro digestion procedure coupled with the use of a human intestinal Caco-2 cell model. Fresh, frozen, and canned spinach were analyzed uncooked and after boiling or microwave cooking. Lutein content from the uncooked and cooked digested food (digestate) and appropriate micelles was determined. Micellarized lutein from the spinach samples was adjusted to 0.1 micromol/L and added to Caco-2 cells. Cellular uptake and secretion (cellular transport) of lutein were determined. Our results showed that digestate obtained from uncooked canned spinach had greater lutein content (P canned spinach digestate and micelles compared with their uncooked counterparts. Interestingly, there were no differences in the micellarization of lutein between the cooking and processing methods. Cellular transport of lutein was greater from uncooked spinach micelles compared with boiled or microwave-cooked spinach. To conclude, although the lutein content of digesta and micelles may have been modified, its micellarization was not significantly affected by any of the cooking or processing methods tested. In general, cellular transport of lutein was greatest in uncooked spinach irrespective of whether the spinach was fresh, frozen, or canned.

  5. Comparative effects of sub-stimulating concentrations of non-human versus human Luteinizing Hormones (LH) or chorionic gonadotropins (CG) on adenylate cyclase activation by forskolin in MLTC cells.

    Science.gov (United States)

    Nguyen, Thi-Mong Diep; Filliatreau, Laura; Klett, Danièle; Combarnous, Yves

    2018-05-15

    We have compared various Luteinizing Hormone (LH) and Chorionic Gonadotropin (CG) preparations from non-human and human species in their ability to synergize with 10 µM forskolin (FSK) for cyclic AMP intracellular accumulation, in MLTC cells. LH from rat pituitary as well as various isoforms of pituitary ovine, bovine, porcine, equine and human LHs and equine and human CG were studied. In addition, recombinant human LH and CG were also compared with the natural human and non-human hormones. Sub-stimulating concentrations of all LHs and CGs (2-100 pM) were found to stimulate cyclic AMP accumulation in MLTC cells in the presence of an also non-stimulating FSK concentration (10 µM). Like rat LH, the most homologous available hormone for mouse MLTC cells, all non-human LHs and CG exhibit a strong potentiating effect on FSK response. The human, natural and recombinant hLH and hCG also do so but in addition, they were found to elicit a permissive effect on FSK stimulation. Indeed, when incubated alone with MLTC cells at non-stimulating concentrations (2-70 pM) hLH and hCG permit, after being removed, a dose-dependent cyclic AMP accumulation with 10 µM FSK. Our data show a clearcut difference between human LH and CG compared to their non-human counterparts on MLTC cells adenylate cyclase activity control. This points out the risk of using hCG as a reference ligand for LHR in studies using non-human cells. Copyright © 2018 Elsevier Inc. All rights reserved.

  6. Effects of nitrogen source availability and bioreactor operating strategies on lutein production with Scenedesmus obliquus FSP-3.

    Science.gov (United States)

    Ho, Shih-Hsin; Xie, Youping; Chan, Ming-Chang; Liu, Chen-Chun; Chen, Chun-Yen; Lee, Duu-Jong; Huang, Chieh-Chen; Chang, Jo-Shu

    2015-05-01

    In this study, the effects of the type and concentration of nitrogen sources on the cell growth and lutein content of an isolated microalga Scenedesmus obliquus FSP-3 were investigated. With batch culture, the highest lutein content (4.61 mg/g) and lutein productivity (4.35 mg/L/day) were obtained when using 8.0 mM calcium nitrate as the nitrogen source. With this best nitrogen source condition, the microalgae cultivation was performed using two bioreactor strategies (namely, semi-continuous and two-stage operations) to further enhance the lutein content and productivity. Using semi-continuous operation with a 10% medium replacement ratio could obtain the highest biomass productivity (1304.8 mg/L/day) and lutein productivity (6.01 mg/L/day). This performance is better than most related studies. Copyright © 2014 Elsevier Ltd. All rights reserved.

  7. Lutein bioavailability from lutein ester-fortified fermented milk: in vivo and in vitro study.

    Science.gov (United States)

    Granado-Lorencio, Fernando; Herrero-Barbudo, Carmer; Olmedilla-Alonso, Begoña; Blanco-Navarro, Inmaculada; Pérez-Sacristán, Belén

    2010-02-01

    We assessed the bioavailability of lutein from lutein-fortified fermented milk using in vivo and in vitro approaches. Twenty-four volunteers were randomized to take lutein-fortified fermented milk at two levels of fortification. Single-dose bioavailability study (2x100 ml, ca. 8 or 16 mg of lutein) was performed using a three-point approach (baseline, 3.5 and 6.5 h). Multiple-dose study consisted of consuming one serving/day (ca. 4 or 8 mg/100 ml) for 14 days. Blood samples for biochemical, hematological and lutein analysis were drawn at baseline, Day 7 and Day 14. In vitro bioaccessibility was assessed by a static gastrointestinal digestion model. Lutein content, in vitro ester hydrolysis and micellarization, and lutein concentrations achieved in serum were analyzed by HPLC. In vivo, post-prandial response was higher using the high content fermented milk, but the percentage of absorption was not different according to the dose consumed. Net increments at Day 7 and Day 14 were significantly higher on consuming the high-dose milk as well. In vitro, lutein ester hydrolysis was incomplete regardless of the amount initially present. Free lutein released was higher using the high-dose fermented milk, but the percentage of hydrolysis was similar at both levels of fortification. In the micellar phase, the percentage of free and total lutein was not different according to the dose. Our results support the suitability of the fermented milk as a carrier of lutein esters and an in vivo dose-dependent effect upon regular consumption and suggest the usefulness of in vitro models to provide relevant information to predict in vivo responses. Copyright 2010 Elsevier Inc. All rights reserved.

  8. Process analysis and modeling of a single-step lutein extraction method for wet microalgae.

    Science.gov (United States)

    Gong, Mengyue; Wang, Yuruihan; Bassi, Amarjeet

    2017-11-01

    Lutein is a commercial carotenoid with potential health benefits. Microalgae are alternative sources for the lutein production in comparison to conventional approaches using marigold flowers. In this study, a process analysis of a single-step simultaneous extraction, saponification, and primary purification process for free lutein production from wet microalgae biomass was carried out. The feasibility of binary solvent mixtures for wet biomass extraction was successfully demonstrated, and the extraction kinetics of lutein from chloroplast in microalgae were first evaluated. The effects of types of organic solvent, solvent polarity, cell disruption method, and alkali and solvent usage on lutein yields were examined. A mathematical model based on Fick's second law of diffusion was applied to model the experimental data. The mass transfer coefficients were used to estimate the extraction rates. The extraction rate was found more significantly related with alkali ratio to solvent than to biomass. The best conditions for extraction efficiency were found to be pre-treatment with ultrasonication at 0.5 s working cycle per second, react 0.5 h in 0.27 L/g solvent to biomass ratio, and 1:3 ether/ethanol (v/v) with 1.25 g KOH/L. The entire process can be controlled within 1 h and yield over 8 mg/g lutein, which is more economical for scale-up.

  9. Central hypogonadism due to a giant, "silent" FSH-secreting, atypical pituitary adenoma: effects of adenoma dissection and short-term Leydig cell stimulation by luteinizing hormone (LH) and human chorionic gonadotropin (hCG).

    Science.gov (United States)

    Santi, Daniele; Spaggiari, Giorgia; Casarini, Livio; Fanelli, Flaminia; Mezzullo, Marco; Pagotto, Uberto; Granata, Antonio R M; Carani, Cesare; Simoni, Manuela

    2017-06-01

    We present a case report of an atypical giant pituitary adenoma secreting follicle-stimulating hormone (FSH). A 55-year-old patient presented for erectile dysfunction, loss of libido and fatigue. The biochemical evaluation showed very high FSH serum levels in the presence of central hypogonadism. Neither testicular enlargement nor increased sperm count was observed, thus a secretion of FSH with reduced biological activity was supposed. The histological examination after neuro-surgery showed an atypical pituitary adenoma with FSH-positive cells. Hypogonadism persisted and semen analyses impaired until azoospermia in conjunction with the reduction in FSH levels suggesting that, at least in part, this gonadotropin should be biologically active. Thus, we hypothesized a concomitant primary testicular insufficiency. The patient underwent short-term treatment trials with low doses of either recombinant luteinizing hormone (LH) or human chorionic gonadotropin (hCG) in three consecutive treatment schemes, showing an equal efficacy in stimulating testosterone (T) increase. This is the first case of atypical, giant FSH-secreting pituitary adenoma with high FSH serum levels without signs of testicular hyperstimulation, in presence of hypogonadism with plausible combined primary and secondary etiology. Hypophysectomized patients may represent a good model to assess both pharmacodynamics and effective dose of LH and hCG in the male.

  10. 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...

  11. Effects of feeding lutein on production performance, antioxidative status, and milk quality of high-yielding dairy cows.

    Science.gov (United States)

    Xu, C Z; Wang, H F; Yang, J Y; Wang, J H; Duan, Z Y; Wang, C; Liu, J X; Lao, Y

    2014-11-01

    This experiment was conducted to determine the influences of supplementing different levels of an additive containing lutein in the diet of Chinese Holstein lactating cows on production performance, antioxidative plasma metabolites, and milk quality. This study was performed on 60 multiparous Holstein dairy cows in peak lactation. The cows were randomly allocated to 1 of 4 homogeneous treatments, with lutein preparation (extracted from marigolds; effective lutein content was 2%) added at levels of 0, 100, 150, and 200 g/d per head, with the actual available amounts being 0, 2, 3, and 4 g of lutein/d per head, respectively. The experiment lasted for 13 wk, with the first week for adaptation. Milk yield and milk compositions were recorded weekly, and milk concentrations of lutein, dry matter intake, and antioxidative blood index were analyzed in the first, fourth, seventh, and thirteenth week of the study. The results showed that adding lutein in the diet had no effect on dry matter intake compared with the control group; however, it slowed down the trend of decline in milk yield, and had a linear incremental effect on milk yield with increasing concentration of lutein. Dietary lutein tended to quadratically increase the percentage of milk fat, and linearly increased milk lactose concentration, with the highest value when treated at 200 g of lutein preparation/d per head, and decreased somatic cell count, with the lowest values when treated with 150 and 200 g of lutein preparation/d per head. The concentration of lutein in milk linearly increased with the incorporation of the additive, with a value of 0.59, 0.70, 1.20, and 1.50 μg/100mL when treated with 0, 100, 150, and 200 g/d, respectively. Total plasma antioxidant capacity tended to linearly increase in cows fed lutein preparation, whereas plasma superoxide dismutase and glutathione peroxidase activities did not differ significantly. In conclusion, addition of lutein in the diet could improve the production

  12. 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...

  13. Association between lutein intake and lutein concentrations in human milk samples from lactating mothers in South Korea.

    Science.gov (United States)

    Kim, Hyesook; Yi, Hyunju; Jung, Ji A; Chang, Namsoo

    2018-02-01

    This study aimed to determine the lutein content of breast milk and its association with maternal lutein intake among lactating mothers in South Korea. Milk samples were obtained from 98 healthy lactating women (mean age; 32.5 ± 3.5 years). Dietary intake data were collected by a food record method for three consecutive days. Maternal lutein intake was estimated by using the lutein database. Lutein concentrations in human milk were analyzed using a high-performance liquid chromatography-ultraviolet detection method. The mean values of the daily lutein intakes and breast milk lutein concentrations in lactating mothers were 4.70 ± 3.11 mg/day (median 3.87) and 3.50 ± 3.71 µg/dl (median 2.45), respectively. Breast milk lutein concentrations were positively associated with the dietary lutein intake of lactating mothers after adjustment for lactating women's age, BMI, dietary energy intake, type of breastfeeding, and infants' age (β = 0.3629, P = 0.0056). Considering that lutein in milk can be associated with dietary lutein intake, knowledge about infant requirement is needed to define the adequate lutein levels in human milk.

  14. Bioavailability of lutein from a lutein-enriched egg-yolk beverage and its dried re-suspended versions

    NARCIS (Netherlands)

    Bunger, M.; Quataert, M.C.J.; Kamps, L.M.; Versloot, P.; Hulshof, P.J.M.; Togtema, K.A.; Amerongen, van A.; Mensink, M.R.

    2014-01-01

    Drying a fresh lutein-enriched egg-yolk beverage would extend its shelf life, however, functional properties should not be affected. It was investigated whether consumption of a dried beverage containing lutein-enriched egg-yolk significantly increases serum lutein. One-hundred healthy young

  15. Synaptology of luteinizing hormone-releasing hormone (LHRH)-immunoreactive cells in the nervus terminalis of the gray short-tailed opossum (Monodelphis domestica).

    Science.gov (United States)

    Zheng, L M; Pfaff, D W; Schwanzel-Fukuda, M

    1990-05-08

    Light and electron microscopic immunocytochemistry were used to examine the structure of LHRH neurons and fibers in the nervus terminalis of the gray short-tailed opossum (Monodelphis domestica). LHRH-immunoreactive neurons and fibers form a loose plexus within the fascicular network of the ganglion terminale on the median surface of the olfactory bulb. There are at least two populations of LHRH-immunoreactive neurons within the network of the ganglion terminale: fusiform and round neurons similar to those described in the forebrain. At the ultrastructural level, axosomatic and axodendritic contacts were seen between LHRH-immunoreactive and nonimmunoreactive elements in the ganglion terminale. These contacts were classified as 1) synaptic input, with asymmetric synapses seen between a nonimmunoreactive axon terminal and a LHRH-immunoreactive cell body or a nonimmunoreactive axon terminal and a LHRH-immunoreactive dendritic process. 2) synaptic output, with symmetric synapses seen between LHRH-immunoreactive and nonimmunoreactive processes. This study is the first systematic examination of the ultrastructure of the LHRH-immunoreactive neurons and their synaptic contacts in the nervus terminalis. The possible integrative roles for this LHRH-immunoreactive system are discussed.

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

    Science.gov (United States)

    Vishwanathan, Rohini; Neuringer, Martha; Snodderly, D. Max; Schalch, Wolfgang; Johnson, Elizabeth J.

    2013-01-01

    Objectives 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 tissue. The objective of the study was to evaluate the relationship between retinal and brain levels of these xanthophylls in non-human primates. Methods Study animals included rhesus monkeys reared on diets devoid of xanthophylls that were subsequently fed pure lutein or pure zeaxanthin (both at 3.9 μmol/kg*d, n=6/group) and normal rhesus monkeys fed a stock diet (0.26 μmol/kg*d lutein and 0.24 μmol/kg*d zeaxanthin, n=5). Retina (4 mm macular punch, 4-8 mm annulus and periphery) and brain tissue (cerebellum, frontal cortex, occipital cortex and pons) from the same animals were analyzed by reverse phase HPLC. Results Lutein in the macula and annulus were significantly related to lutein levels in the cerebellum, occipital cortex and pons, both in bivariate analysis and after adjusting for age, sex and n–3 fatty acid status. In the frontal cortex the relationship was marginally significant. Macular zeaxanthin was significantly related to zeaxanthin in the cerebellum and frontal cortex, while the relationship was marginally significant in the occipital cortex and pons in a bivariate model. Discussion An integrated measure of total macular pigment optical density, which can be measured noninvasively, has the potential to be used as a biomarker to assess brain lutein and zeaxanthin status. PMID:22780947

  17. Lutein Supplementation Increases Breast Milk and Plasma Lutein Concentrations in Lactating Women and Infant Plasma Concentrations but Does Not Affect Other Carotenoids 1 2 3

    OpenAIRE

    Sherry, Christina L.; Oliver, Jeffery S.; Renzi, Lisa M.; Marriage, Barbara J.

    2014-01-01

    Lutein is a carotenoid that varies in breast milk depending on maternal intake. Data are lacking with regard to the effect of dietary lutein supplementation on breast milk lutein concentration during lactation and subsequent plasma lutein concentration in breast-fed infants. This study was conducted to determine the impact of lutein supplementation in the breast milk and plasma of lactating women and in the plasma of breast-fed infants 2–3 mo postpartum. Lutein is the dominant carotenoid in t...

  18. Luteinized fat in Krukenberg tumor: MR findings

    International Nuclear Information System (INIS)

    Jeong, Yong Yeon; Kang, Heoung Keun; Seo, Jeong Jin; Nam, Jong Hee

    2002-01-01

    To our knowledge, there is no description of the fat-containing Krukenberg tumor. We report on a case of Krukenberg tumor associated with luteinized fat, which showed hyperintensity on T1-weighted MR image. The diagnosis was surgically confirmed. Hyperintense portion of the Krukenberg tumor on T1-weighted image showed diminished signal intensity on fat-saturated, T1-weighted images. Krukenberg tumor should be considered in the differential diagnosis of ovarian masses when fat signal is seen. (orig.)

  19. Simultaneous radioimmunoassay for luteinizing hormone and prolactin

    International Nuclear Information System (INIS)

    Steele, M.K.; Deschepper, C.F.

    1985-01-01

    A combined radioimmunoassay (RIA) for the measurement of the anterior pituitary proteins luteinizing hormone (LH) and prolactin (PRL) is described and compared with individual RIAs for these hormones. The standard curves and the sample values for LH and PRL were identical when determined in a combined or in an individual RIA. This technique may prove useful to a number of laboratories where it is desirable to determine levels of more than one hormone in limited sample volumes

  20. Identification of StARD3 as a lutein-binding protein in the macula of the primate retina.

    Science.gov (United States)

    Li, Binxing; Vachali, Preejith; Frederick, Jeanne M; Bernstein, Paul S

    2011-04-05

    Lutein, zeaxanthin, and their metabolites are the xanthophyll carotenoids that form the macular pigment of the human retina. Epidemiological evidence suggests that high levels of these carotenoids in the diet, serum, and macula are associated with a decreased risk of age-related macular degeneration (AMD), and the AREDS2 study is prospectively testing this hypothesis. Understanding the biochemical mechanisms underlying the selective uptakes of lutein and zeaxanthin into the human macula may provide important insights into the physiology of the human macula in health and disease. GSTP1 is the macular zeaxanthin-binding protein, but the identity of the human macular lutein-binding protein has remained elusive. Prior identification of the silkworm lutein-binding protein (CBP) as a member of the steroidogenic acute regulatory domain (StARD) protein family and selective labeling of monkey photoreceptor inner segments with an anti-CBP antibody provided an important clue for identifying the primate retina lutein-binding protein. The homology of CBP with all 15 human StARD proteins was analyzed using database searches, Western blotting, and immunohistochemistry, and we here provide evidence to identify StARD3 (also known as MLN64) as a human retinal lutein-binding protein. Antibody to StARD3, N-62 StAR, localizes to all neurons of monkey macular retina and especially cone inner segments and axons, but does not colocalize with the Müller cell marker, glutamine synthetase. Further, recombinant StARD3 selectively binds lutein with high affinity (K(D) = 0.45 μM) when assessed by surface plasmon resonance (SPR) binding assays. Our results demonstrate previously unrecognized, specific interactions of StARD3 with lutein and provide novel avenues for exploring its roles in human macular physiology and disease.

  1. 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

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

    International Nuclear Information System (INIS)

    Bajusz, S.; Janaky, T.; Csernus, V.J.; Bokser, L.; Fekete, M.; Srkalovic, G.; Redding, T.W.; Schally, A.V.

    1989-01-01

    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

  3. 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; Martínez-Jerónimo, Fernando; Flores-Ortíz, Cesar Mateo

    2017-11-20

    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 (Y x/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.

  4. Effect of type of TAG fatty acids on lutein and zeaxanthin bioavailability.

    Science.gov (United States)

    Gleize, Béatrice; Tourniaire, Franck; Depezay, Laurence; Bott, Romain; Nowicki, Marion; Albino, Lionel; Lairon, Denis; Kesse-Guyot, Emmanuelle; Galan, Pilar; Hercberg, Serge; Borel, Patrick

    2013-07-14

    The xanthophylls lutein and zeaxanthin probably play a role in visual function and may participate in the prevention of age-related eye diseases. Although a minimum amount of TAG is required for an optimal bioavailability of these carotenoids, the effect of the type of TAG fatty acids (FA) is less clear. The aim was to assess the effect of the type of TAG FA on bioavailability of these xanthophylls. A total of three complementary models were used: an in vitro digestion model to study bioaccessibility, Caco-2 cells to study uptake efficiency and orally administered rats to study in vivo bioavailability. Results showed that lutein and zeaxanthin bioaccessibility was greater (about 20-30 %, Pxanthophyll uptake by Caco-2 cells, but some compounds present in natural oils significantly affected xanthophyll uptake. Oral administration of rats with spinach and butter over 3 d led to a higher fasting plasma lutein concentration than oral administration with olive or fish oils. In conclusion, dietary fats rich in SFA lead to a higher bioavailability of lutein and zeaxanthin, as compared with fats rich in MUFA and PUFA. This is due partly to the higher bioaccessibility of these xanthophylls in the smaller mixed micelles produced when SFA are incorporated into mixed micelles.

  5. 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.

  6. Oxidative stability of yogurt with added lutein dye.

    Science.gov (United States)

    Domingos, L D; Xavier, A A O; Mercadante, A Z; Petenate, A J; Jorge, R A; Viotto, W H

    2014-02-01

    This study evaluated the effect of adding lutein dye on the oxidative stability of yogurt during 35 d of refrigerated storage, in the presence and absence of light. Yogurts manufactured without and with the equivalent of 1.5mg of lutein in 120 g of the final product were characterized for their total carotenoid and riboflavin contents, and the behaviors of both riboflavin and lutein were monitored during storage. A decrease in riboflavin content occurred, with concurrent appearance of its derived-oxidation products in the yogurts without added lutein and exposed to light during storage. The yogurts with added lutein dye showed constant lutein and riboflavin contents throughout storage both for the samples stored under light and for those stored in the dark. Yogurts (120 g) with the addition of 0.5, 1.5, and 2.5mg of lutein dye were evaluated for their sensory acceptance, and the statistical analysis showed no differences between the samples for the attributes of aroma and flavor. These results indicate that the added lutein remained stable throughout the storage period and conferred protection for the riboflavin against photooxidation, preserving the quality of the yogurts. Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  7. Combined Effect of Cameo2 and CBP on the Cellular Uptake of Lutein in the Silkworm, Bombyx mori

    Science.gov (United States)

    Dong, Xiao-Long; Chai, Chun-Li; Pan, Cai-Xia; Tang, Hui; Chen, Yan-Hong; Dai, Fang-Yin; Pan, Min-Hui; Lu, Cheng

    2014-01-01

    Formation of yellow-red color cocoons in the silkworm, Bombyx mori, occurs as the result of the selective delivery of carotenoids from the midgut to the silk gland via the hemolymph. This process of pigment transport is thought to be mediated by specific cellular carotenoids carrier proteins. Previous studies indicated that two proteins, Cameo2 and CBP, are associated with the selective transport of lutein from the midgut into the silk gland in Bombyx mori. However, the exact roles of Cameo2 and CBP during the uptake and transport of carotenoids are still unknown. In this study, we investigated the respective contributions of these two proteins to lutein and β-carotene transport in Bombyx mori as well as commercial cell-line. We found that tissues, expressed both Cameo2 and CBP, accumulate lutein. Cells, co-expressed Cameo2 and CBP, absorb 2 fold more lutein (PBombyx mori. Cameo2 and CBP, as the membrane protein and the cytosol protein, respectively, have the combined effect to facilitate the cellular uptake of lutein. PMID:24475153

  8. Lutein and preterm infants with decreased concentrations of brain carotenoids.

    Science.gov (United States)

    Vishwanathan, Rohini; Kuchan, Matthew J; Sen, Sarbattama; Johnson, Elizabeth J

    2014-11-01

    Lutein and zeaxanthin are dietary carotenoids that may influence visual and cognitive development. The objective of this study was to provide the first data on distribution of carotenoids in the infant brain and compare concentrations in preterm and term infants. Voluntarily donated brain tissues from 30 infants who died during the first 1.5 years of life were obtained from the Eunice Kennedy Shriver National Institute of Child Health and Human Development Brain and Tissue Bank. Tissues (hippocampus and prefrontal, frontal, auditory, and occipital cortices) were extracted using standard lipid extraction procedures and analyzed using reverse-phase high-pressure liquid chromatography. Lutein, zeaxanthin, cryptoxanthin, and β-carotene were the major carotenoids found in the infant brain tissues. Lutein was the predominant carotenoid accounting for 59% of total carotenoids. Preterm infants (n = 8) had significantly lower concentrations of lutein, zeaxanthin, and cryptoxanthin in their brain compared with term infants (n = 22) despite similarity in postmenstrual age. Among formula-fed infants, preterm infants (n = 3) had lower concentrations of lutein and zeaxanthin compared with term infants (n = 5). Brain lutein concentrations were not different between breast milk-fed (n = 3) and formula-fed (n = 5) term decedents. In contrast, term decedents with measurable brain cryptoxanthin, a carotenoid that is inherently low in formula, had higher brain lutein, suggesting that the type of feeding is an important determinant of brain lutein concentrations. These data reveal preferential accumulation and maintenance of lutein in the infant brain despite underrepresentation in the typical infant diet. Further investigation on the impact of lutein on neural development in preterm infants is warranted.

  9. 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.

  10. Luteinizing hormone signaling phosphorylates and activates the cyclic GMP phosphodiesterase PDE5 in mouse ovarian follicles, contributing an additional component to the hormonally induced decrease in cyclic GMP that reinitiates meiosis.

    Science.gov (United States)

    Egbert, Jeremy R; Yee, Siu-Pok; Jaffe, Laurinda A

    2018-03-01

    Prior to birth, oocytes within mammalian ovarian follicles initiate meiosis, but then arrest in prophase until puberty, when with each reproductive cycle, one or more follicles are stimulated by luteinizing hormone (LH) to resume meiosis in preparation for fertilization. Within preovulatory follicles, granulosa cells produce high levels of cGMP, which diffuses into the oocyte to maintain meiotic arrest. LH signaling restarts meiosis by rapidly lowering the levels of cGMP in the follicle and oocyte. Part of this decrease is mediated by the dephosphorylation and inactivation the NPR2 guanylyl cyclase in response to LH, but the mechanism for the remainder of the cGMP decrease is unknown. At least one cGMP phosphodiesterase, PDE5, is activated by LH signaling, which would contribute to lowering cGMP. PDE5 exhibits increased cGMP-hydrolytic activity when phosphorylated on serine 92, and we recently demonstrated that LH signaling phosphorylates PDE5 on this serine and increases its activity in rat follicles. To test the extent to which this mechanism contributes to the cGMP decrease that restarts meiosis, we generated a mouse line in which serine 92 was mutated to alanine (Pde5-S92A), such that it cannot be phosphorylated. Here we show that PDE5 phosphorylation is required for the LH-induced increase in cGMP-hydrolytic activity, but that this increase has only a modest effect on the LH-induced cGMP decrease in mouse follicles, and does not affect the timing of meiotic resumption. Though we show that the activation of PDE5 is among the mechanisms contributing to the cGMP decrease, these results suggest that another cGMP phosphodiesterase is also activated by LH signaling. Copyright © 2018 Elsevier Inc. All rights reserved.

  11. Compared with Powdered Lutein, a Lutein Nanoemulsion Increases Plasma and Liver Lutein, Protects against Hepatic Steatosis, and Affects Lipoprotein Metabolism in Guinea Pigs.

    Science.gov (United States)

    Murillo, Ana Gabriela; Aguilar, David; Norris, Gregory H; DiMarco, Diana M; Missimer, Amanda; Hu, Siqi; Smyth, Joan A; Gannon, Sarah; Blesso, Christopher N; Luo, Yangchao; Fernandez, Maria Luz

    2016-10-01

    It is not clear how oil-in-water nanoemulsions of lutein may affect bioavailability and consequently alter lipoprotein metabolism, oxidative stress, and inflammation. The bioavailability as well as effects of a powdered lutein (PL) and an oil-in-water lutein nanoemulsion (NANO; particle size: 254.2 nm; polydispersity index: 0.29; and ζ-potential: -65 mV) on metabolic variables in liver, plasma, and adipose tissue in a guinea pig model of hepatic steatosis were evaluated. Twenty-four 2-mo-old male Hartley guinea pigs, weighing 200-300 g (n = 8/group), were fed diets containing 0.25 g cholesterol/100 g to induce liver injury for the duration of the study. They were allocated to control (0 mg lutein), PL (3.5 mg/d), or NANO (3.5 mg/d) groups. After 6 wk, plasma, liver, and adipose tissue were collected for determination of lutein, plasma lipids, tissue cholesterol, and inflammatory cytokines. The NANO group had 2-fold higher concentrations of lutein in plasma (P guinea pigs. © 2016 American Society for Nutrition.

  12. Radioimmunoassay of luteinizing hormone in hypothyroidism

    International Nuclear Information System (INIS)

    Sobieszczyk, S.

    1975-01-01

    Radioimmunoassay of luteinizing hormone was performed in 18 women with primary hypothyroidism and 15 women with secondary hypothyroidism. The results of determinations were compared with LH values found in healthy women at reproductive age and after menopause. It was observed that in primary hypothyroidism the level of LH is normal, in young women it was from 6 to 25 m IU/ml, while in the postmenopausal period it increased to 70 to 200 m IU/ml. In secondary hypothyroidism due to pituitary hypofunction the LH level is undetectable or lies in the range of lowest values observed in healthy subjects, not exceeding 8 m IU/ml. Determinations of serum LH may be useful for differential diagnosis of primary and secondary hypothyroidism. (author)

  13. Role of lutein and zeaxanthin in visual and cognitive function throughout the lifespan

    Science.gov (United States)

    The relationship between lutein and zeaxanthin and visual and cognitive health throughout the lifespan is compelling. There is a variety of evidence to support a role for lutein and zeaxanthin in vision. Lutein's role in cognition has only recently been considered. Lutein and its isomer, zeaxanthin,...

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

    OpenAIRE

    Ramirez Maria

    2016-01-01

    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 c...

  15. 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

  16. Effect of lutein intervention on visual function in patients with early age-related macular degeneration

    Directory of Open Access Journals (Sweden)

    Chan Li

    2017-11-01

    Full Text Available AIM: To study the effect of lutein intervention on visual function of patients with early age-related macular degeneration(AMD. METHODS: Totally 200 early AMD patients were divided into lutein intervention group(20mg/dand placebo group by a randomized, double-blind, placebo-controlled trail. Questionnaire investigation, serum lutein concentration and visual function were conducted at baseline, 12, 24, 36 and 48wk respectively. RESULTS: The serum lutein concentration in lutein intervention group was higher than the baseline(PPPPP>0.05. CONCLUSION: Lutein intervention can improve the visual function of patients with early AMD.

  17. Complexes of lutein with bovine and caprine caseins and their impact on lutein chemical stability in emulsion systems: Effect of arabinogalactan.

    Science.gov (United States)

    Mora-Gutierrez, A; Attaie, R; Núñez de González, M T; Jung, Y; Woldesenbet, S; Marquez, S A

    2018-01-01

    Lutein is an important xanthophyll carotenoid with many benefits to human health. Factors affecting the application of lutein as a functional ingredient in low-fat dairy-like beverages (pH 6.0-7.0) are not well understood. The interactions of bovine and caprine caseins with hydrophobic lutein were studied using UV/visible spectroscopy as well as fluorescence. Our studies confirmed that the aqueous solubility of lutein is improved after binding with bovine and caprine caseins. The rates of lutein solubilization by the binding to bovine and caprine caseins were as follows: caprine α S1 -II-casein 34%, caprine α S1 -I-casein 10%, and bovine casein 7% at 100 μM lutein. Fluorescence of the protein was quenched on binding supporting complex formation. The fluorescence experiments showed that the binding involves tryptophan residues and some nonspecific interactions. Scatchard plots of lutein binding to the caseins demonstrated competitive binding between the caseins and their sites of interaction with lutein. Competition experiments suggest that caprine α S1 -II casein will bind a larger number of lutein molecules with higher affinity than other caseins. The chemical stability of lutein was largely dependent on casein type and significant increases occurred in the chemical stability of lutein with the following pattern: caprine α S1 -II-casein > caprine α S1 -I-casein > bovine casein. Addition of arabinogalactan to lutein-enriched emulsions increases the chemical stability of lutein-casein complexes during storage under accelerated photo-oxidation conditions at 25°C. Therefore, caprine α S1 -II-casein alone and in combination with arabinogalactan can have important applications in the beverage industry as carrier of this xanthophyll carotenoid (lutein). Copyright © 2018 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  18. Lutein Supplementation Increases Breast Milk and Plasma Lutein Concentrations in Lactating Women and Infant Plasma Concentrations but Does Not Affect Other Carotenoids123

    Science.gov (United States)

    Sherry, Christina L.; Oliver, Jeffery S.; Renzi, Lisa M.; Marriage, Barbara J.

    2014-01-01

    Lutein is a carotenoid that varies in breast milk depending on maternal intake. Data are lacking with regard to the effect of dietary lutein supplementation on breast milk lutein concentration during lactation and subsequent plasma lutein concentration in breast-fed infants. This study was conducted to determine the impact of lutein supplementation in the breast milk and plasma of lactating women and in the plasma of breast-fed infants 2–3 mo postpartum. Lutein is the dominant carotenoid in the infant brain and the major carotenoid found in the retina of the eye. Eighty-nine lactating women 4–6 wk postpartum were randomly assigned to be administered either 0 mg/d of lutein (placebo), 6 mg/d of lutein (low-dose), or 12 mg/d of lutein (high-dose). The supplements were consumed for 6 wk while mothers followed their usual diets. Breast milk carotenoids were measured weekly by HPLC, and maternal plasma carotenoid concentrations were measured at the beginning and end of the study. Infant plasma carotenoid concentrations were assessed at the end of the study. No significant differences were found between dietary lutein + zeaxanthin intake and carotenoid concentrations in breast milk and plasma or body mass index at baseline. Total lutein + zeaxanthin concentrations were greater in the low- and high-dose–supplemented groups than in the placebo group in breast milk (140% and 250%, respectively; P Lutein supplementation did not affect other carotenoids in lactating women or their infants. Lactating women are highly responsive to lutein supplementation, which affects plasma lutein concentrations in the infant. This trial was registered at clinicaltrials.gov as NCT01747668. PMID:24899160

  19. Lutein supplementation increases breast milk and plasma lutein concentrations in lactating women and infant plasma concentrations but does not affect other carotenoids.

    Science.gov (United States)

    Sherry, Christina L; Oliver, Jeffery S; Renzi, Lisa M; Marriage, Barbara J

    2014-08-01

    Lutein is a carotenoid that varies in breast milk depending on maternal intake. Data are lacking with regard to the effect of dietary lutein supplementation on breast milk lutein concentration during lactation and subsequent plasma lutein concentration in breast-fed infants. This study was conducted to determine the impact of lutein supplementation in the breast milk and plasma of lactating women and in the plasma of breast-fed infants 2-3 mo postpartum. Lutein is the dominant carotenoid in the infant brain and the major carotenoid found in the retina of the eye. Eighty-nine lactating women 4-6 wk postpartum were randomly assigned to be administered either 0 mg/d of lutein (placebo), 6 mg/d of lutein (low-dose), or 12 mg/d of lutein (high-dose). The supplements were consumed for 6 wk while mothers followed their usual diets. Breast milk carotenoids were measured weekly by HPLC, and maternal plasma carotenoid concentrations were measured at the beginning and end of the study. Infant plasma carotenoid concentrations were assessed at the end of the study. No significant differences were found between dietary lutein + zeaxanthin intake and carotenoid concentrations in breast milk and plasma or body mass index at baseline. Total lutein + zeaxanthin concentrations were greater in the low- and high-dose-supplemented groups than in the placebo group in breast milk (140% and 250%, respectively; P Lutein supplementation did not affect other carotenoids in lactating women or their infants. Lactating women are highly responsive to lutein supplementation, which affects plasma lutein concentrations in the infant. This trial was registered at clinicaltrials.gov as NCT01747668. © 2014 American Society for Nutrition.

  20. Lutein and Zeaxanthin Isomers in Eye Health and Disease.

    Science.gov (United States)

    Mares, Julie

    2016-07-17

    Current evidence suggests lutein and its isomers play important roles in ocular development in utero and throughout the life span, in vision performance in young and later adulthood, and in lowering risk for the development of common age-related eye diseases in older age. These xanthophyll (oxygen-containing) carotenoids are found in a wide variety of vegetables and fruits, and they are present in especially high concentrations in leafy green vegetables. Additionally, egg yolks and human milk appear to be bioavailable sources. The prevalence of lutein, zeaxanthin, and meso-zeaxanthin in supplements is increasing. Setting optimal and safe ranges of intake requires additional research, particularly in pregnant and lactating women. Accumulating evidence about variable interindividual response to dietary intake of these carotenoids, based on genetic or metabolic influences, suggests that there may be subgroups that benefit from higher levels of intake and/or alternate strategies to improve lutein and zeaxanthin status.

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

    Science.gov (United States)

    Bajusz, S; Janaky, T; Csernus, V J; Bokser, L; Fekete, M; Srkalovic, G; Redding, T W; Schally, A V

    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. For instance, SB-40, a PtCl2-containing metallopeptide in which platinum is coordinated to an N epsilon-(DL-2,3-diaminopropionyl)-D-lysine residue [D-Lys(DL-A2pr] at position 6, showed 50 times higher LH-releasing potency than the native hormone. SB-95, [Ac-D-Nal(2)1,D-Phe(pCl)2, D-Pal(3)2, Arg5,D-Lys[DL-A2pr(Sal2Cu)]6,D-Ala10]LH-RH, where Nal(2) is 3-(2-naphthyl)alanine, Pal(3) is 3-(3-pyridyl)alanine, and copper(II) is coordinated to the salicylideneimino moieties resulting from condensation of salicylaldehyde with D-Lys(DL-A2pr)6, caused 100% inhibition of ovulation at a dose of 3 micrograms in rats. 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 cell lines in vitro (this will be the subject of a separate paper on cytotoxicity evaluation). Such cytostatic metallopeptides could be envisioned as targeted chemotherapeutic agents in cancers that contain receptors for LH-RH-like peptides.

  2. 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.

  3. Evidence for alterations in luteinizing hormone secreted in rhesus monkeys with normal and inadequate luteal phases using radioreceptor and radioimmunoassay

    Energy Technology Data Exchange (ETDEWEB)

    Sakai, C.N.; Channing, C.P.

    1979-05-01

    A radioreceptor assay (RRA) using porcine granulosa cells and (/sup 125/I)hCG was developed and validated for the measurement of LH or CG. The RRA was used in conjunction with a heterologous RIA employing antiserum against ovine LH and (/sup 125/I)ovine LH (RIA) to measure serum LH in the rhesus monkey throughout the menstrual cycle. Discrepancies were found in the measurement of serum LH using RRA and RIA. Measurements of serum LH using RIA were consistently higher than the measurements of serum LH using RRA in serum from adult intact female and male monkeys and hypophysectomized, ovariectomized, and pregnant monkeys.

  4. Improved oral bioavailability for lutein by nanocrystal technology: formulation development, in vitro and in vivo evaluation.

    Science.gov (United States)

    Chang, Daoxiao; Ma, Yanni; Cao, Guoyu; Wang, Jianhuan; Zhang, Xia; Feng, Jun; Wang, Wenping

    2018-08-01

    Lutein is a kind of natural carotenoids possessing many pharmacological effects. The application of lutein was limited mainly due to its low oral bioavailability caused by poor aqueous solubility. Nanocrystal formulation of lutein was developed to improve the oral bioavailability in this study. The nanosuspension was prepared by the anti-solvent precipitation-ultrasonication method and optimized by Box-Behnken design, followed by freeze-drying to obtain lutein nanocrystals. The nanocrystals were characterized on their physical properties, in vitro dissolution and in vivo absorption performance. Lutein nanocrystals showed as tiny spheres with an average particle size of 110.7 nm. The result of diffractograms indicated that the percent crystallinity of lutein was 89.4% in coarse powder and then declined in nanocrystal formulation. The saturated solubility of lutein in water increased from 7.3 μg/ml for coarse powder up to 215.7 μg/ml for lutein nanocrystals. The dissolution rate of lutein nanocrystals was significantly higher than that of coarse powder or the physical mixture. The C max and AUC 0-24 h of lutein nanocrystals after oral administration in rats was 3.24 and 2.28 times higher than those of lutein suspension, respectively. These results indicated that the nanocrystal formulation could significantly enhance the dissolution and absorption of lutein and might be a promising approach for improving its oral bioavailability.

  5. Biocompatible lutein-polymer-lipid nanocapsules: Acute and subacute toxicity and bioavailability in mice

    Energy Technology Data Exchange (ETDEWEB)

    Ranganathan, Arunkumar; Hindupur, Ravi; Vallikannan, Baskaran, E-mail: baskaranv@cftri.res.in

    2016-12-01

    Lutein-poly-(lactic-co-glycolic acid) (PLGA)-phospholipid (PL) nanocapsules were prepared (henceforth referred as lutein nanocapsules) and studied for acute, subacute oral toxicity and bioavailability of lutein in mice. Prior to examining the safety of lutein nanocapsules, particle size, zeta potential, surface morphology and interaction between lutein, PLGA and PL were studied. In acute study, mice were gavaged with a single dose of lutein nanocapsules at 0.1, 1, 10 and 100 mg/kg body weight (BW) and examined for 2 weeks, while in subacute study, daily mice were gavaged with a dose of 1 and 10 mg/kg BW for 4 weeks. Results revealed that mean size and zeta value of lutein nanocapsules were 140 nm and − 44 mV, respectively. Acute and subacute toxicity studies did not show any mortality or treatment related adverse effect in clinical observations, ophthalmic examinations, body and organ weights. No toxicity related findings were observed in hematology, histopathology and other blood and tissue clinical chemistry parameters. In subacute study, no observed adverse effect level (NOAEL) of lutein nanocapsules was found to be at a dose of 10 mg/kg BW. Feeding lutein nanocapsules resulted in a significant (p < 0.01) increase in lutein level in plasma and tissue compared to the control group. Lutein nanocapsules did not cause toxicity in mice. However, human trials are warranted. - Highlights: • Acute and subacute toxicity studies of lutein-PLGA-PL showed no toxicity. • PLGA-PL nanocapsules were safe carriers for oral delivery of lutein. • Oral gavage of lutein-PLGA-PL nanocapsule improves plasma lutein levels.

  6. Pituitary Gonadotropins, Prolactin and Growth Hormone Differentially Regulate AQP1 Expression in the Porcine Ovarian Follicular Cells

    Directory of Open Access Journals (Sweden)

    Mariusz T. Skowronski

    2017-12-01

    Full Text Available The present in vitro study analyzed whether the hormones that affect the ovarian follicular steroidogenesis process also participate in the regulation of AQP1 mRNA and protein expression. Granulosa (Gc and theca cells (Tc of medium and large porcine ovarian follicles were exposed to follicle-stimulating hormone (FSH, luteinizing hormone (LH, prolactin (PRL and growth hormone (GH for 24 h in separated cells and co-cultures of these cells. Real-time PCR, Western blotting, immunofluorescence and volumetric analysis were then performed. Gonadotropins, PRL and GH had a stimulatory impact on AQP1 mRNA and protein expression in Gc and Tc of medium and large ovarian cells. Moreover, swelling assays, in response to a hypotonic environment, demonstrated the functional presence of AQPs in porcine Gc and Tc. Immunofluorescence analysis showed that AQP1 protein was mainly localized in the perinuclear region of the cytoplasm, endosomes and cell membranes of Gc and Tc from medium and large follicles. It seems possible that AQP1 present in Gc and Tc cells may be implicated not only in the regulation of water homeostasis required for follicle development but also in cell proliferation and migration.

  7. Dietary Components Affect the Plasma and Tissue Levels of Lutein in Aged Rats with Lutein Deficiency--A Repeated Gavage and Dietary Study.

    Science.gov (United States)

    Sheshappa, Mamatha Bangera; Ranganathan, Arunkumar; Bhatiwada, Nidhi; Talahalli, Ramprasad Ravichandra; Vallikannan, Baskaran

    2015-10-01

    The aim of this study was to find out the influence of selected dietary components on plasma and tissue response of repeated micellar and dietary lutein in aged rats with lutein deficiency. In repeated (16 d) gavage study, micellar lutein was co-ingested with either phosphatidylcholine (PC), lyso-phosphatidylcholine (lysoPC), β-carotene, dietary fiber or vegetable fat (3% soybean oil). In dietary study, rats were fed (4 wk) semi-synthetic diet either with lutein + PC, lutein + dietary fiber or B. alba (lutein source) + PC. The post-prandial plasma and tissue response of lutein was measured by HPLC. Results showed that micellar fat, PC and lysoPC significantly (P ≤ 0.05) increased the lutein levels in plasma (31.1%, 26.8%, and 34.9%), liver (27.4%, 29.5%, and 8.6%), and eyes (63.5%, 90.2%, and 86%) compared to the control group (group gavaged micelles with no dietary components studied). Similarly, dietary study showed an enhanced plasma, liver, and eye lutein levels by 44.8%, 24.1%, and 42.0% (lutein + PC group) and 51.7%, 39.8%, and 31.7% (B.alba + PC group), respectively compared to control. The activity of antioxidant enzymes in plasma and liver of both the studies were also affected compared to control. Result reveals, that PC enhance the intestinal absorption of both micellar and dietary lutein which is either in free or bound form with food matrices in aged rats with lutein deficiency. Hence, PC at a concentration used in this study can be considered to improve the lutein bioavailability in lutein deficiency. Lutein and zeaxanthin are macular pigments acquired mostly from greens, that play an significant role in protecting vision from Age related macular degeneration (AMD). However, their biological availability is poor and affected by dietary components. This study demonstrates the positive influence of dietary PC and lyso PC in improving intestinal uptake of lutein. Our previous and present finding shows there is a possibility of developing functional

  8. 61 HUGE BENIGN GRANULOSA CELL TUMOUR IN A 61 YEAR ...

    African Journals Online (AJOL)

    Dr. E. P. Gharoro

    peritoneal cavity, huge right ovarian cyst measuring 37cm/29cm as in figure 1a, weighing 8.3 kg with a thick smooth wall without excrescences on surface. ... is released in the blood during pregnancy and is produced in other conditions such as endometriosis, fibroids and diverticulitis. It is useful in monitoring therapy.

  9. Acute pancreatitis following granulosa cell tumor removal in a mare

    Science.gov (United States)

    Gomez, Diego E.; Radtke, Catherine L.; Russell, Lauren A.; Lopez, Alfonso; Wichtel, Maureen W.

    2015-01-01

    Acute pancreatitis is a rare disease in horses and is often associated with gastrointestinal disorders. Accurate diagnosis is challenging due to the presence of nonspecific clinical signs. This case represents the first documentation of acute pancreatitis in a horse following surgery of the reproductive tract. PMID:26483579

  10. Highly potent antagonists of luteinizing hormone-releasing hormone free of edematogenic effects.

    Science.gov (United States)

    Bajusz, S; Kovacs, M; Gazdag, M; Bokser, L; Karashima, T; Csernus, V J; Janaky, T; Guoth, J; Schally, A V

    1988-03-01

    To eliminate the undesirable edematogenic effect of the luteinizing hormone-releasing hormone (LH-RH) antagonists containing basic D amino acids at position 6, exemplified by [Ac-D-Phe(pCl)1,2,D-Trp3,D-Arg6,D-Ala10]LH-RH [Phe(pCl) indicates 4-chlorophenylalanine], analogs with D-ureidoalkyl amino acids such as D-citrulline (D-Cit) or D-homocitrulline (D-Hci) at position 6 were synthesized and tested in several systems in vitro and in vivo. HPLC analysis revealed that the overall hydrophobicity of the D-Cit/D-Hci6 analogs was similar to that of the basic D-Arg6 antagonists. In vitro, most of the analogs completely inhibited LH-RH-mediated luteinizing hormone release in perfused rat pituitary cell systems at an antagonist to LH-RH molar ratio of 5:1. In vivo, the most active peptides, [Ac-D-Nal(2)1,D-Phe(pCl)2,D-Trp3,D-Cit6,D-Ala10]LH-RH [Nal(2) indicates 3-(2-naphthyl)alanine] and its D-Hci6 analog, caused 100% inhibition of ovulation in cycling rats in doses of 3 micrograms and suppressed the luteinizing hormone level in ovariectomized female rats for 47 hr when administered at doses of 25 micrograms. Characteristically, these peptides did not exert any edematogenic effects even at 1.5 mg/kg. These properties of the D-Cit/D-Hci6 antagonists may make them useful clinically.

  11. Effect of ultrasonic waves on the stability of all-trans lutein and its degradation kinetics.

    Science.gov (United States)

    Song, Jiang-Feng; Li, Da-Jing; Pang, Hui-Li; Liu, Chun-Quan

    2015-11-01

    Ultrasound treatment has been widely applied in the extraction of biologically active compounds including carotenoids. However, there are few reports on their effects on the stability of these compounds. In the present study, the stability of all-trans lutein, one of the carotenoids, was investigated under the action of ultrasound. Results showed that ultrasound induced the isomerization of all-trans lutein to its isomers, namely to 13-cis lutein, 13'-cis lutein, 9-cis lutein and 9'-cis lutein as analyzed by HPLC coupled with DAD and LC-MS; and the percentage of the isomerization increased with increasing both ultrasonic frequency and power. The stability of all-trans lutein in dichloromethane was worst among multiple kinds of solvents. Interestingly, the retention rate of all-trans lutein improved as the temperature increased, which runs counter to the Arrhenius law. Under ultrasound irradiation, the degradation mechanism might be different with various temperatures, the degradation of all-trans lutein followed first-order kinetics at 20°C, while second-order kinetics was followed at 30-50°C. As the ultrasonic reaction time prolonged, lutein epoxidation nearly occurred. Those results presented here emphasized that UAE techniques should be carefully used in the extraction of all-trans lutein. Copyright © 2015 Elsevier B.V. All rights reserved.

  12. Lutein-fortified infant formula fed to healthy term infants: evaluation of growth effects and safety.

    Science.gov (United States)

    Capeding, Rosario; Gepanayao, Connie P; Calimon, Nerrisa; Lebumfacil, Jowena; Davis, Anne M; Stouffer, Nicole; Harris, Bruce J

    2010-05-21

    Breast milk contains lutein derived from the mother's diet. This carotenoid is currently not added to infant formula, which has a small and variable lutein content from innate ingredients. This study was conducted to compare the growth of infants fed lutein-fortified infant formula with that of infants fed infant formula without lutein fortification. This 16-week study was prospective, randomized, controlled, and double-blind with parallel groups of healthy term infants fed either control formula (Wyeth S-26 Gold, designated as Gold) or experimental formula (Wyeth S-26 Gold fortified with lutein at 200 mcg/l, designated as Gold+Lutein). Two hundred thirty-two (232) infantslutein-fortified S-26 Gold demonstrated growth equivalent to that of infants fed unfortified lutein formula.

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

    Directory of Open Access Journals (Sweden)

    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.

  14. 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%.

  15. Analogues of luteinizing hormone-releasing hormone containing cytotoxic groups.

    Science.gov (United States)

    Janáky, T; Juhász, A; Bajusz, S; Csernus, V; Srkalovic, G; Bokser, L; Milovanovic, S; Redding, T W; Rékási, Z; Nagy, A

    1992-02-01

    In an attempt to produce better cytotoxic analogues, chemotherapeutic antineoplastic radicals including an alkylating nitrogen mustard derivative of D-phenylalanine (D-melphalan), reactive cyclopropane, anthraquinone derivatives [2-(hydroxymethyl)anthraquinone and the anticancer antibiotic doxorubicin], and an antimetabolite (methotrexate) were coupled to suitably modified agonists and antagonists of luteinizing hormone-releasing hormone (LH-RH). Analogues with D-lysine6 and D-ornithine6 or N epsilon-(2,3-diaminopropionyl)-D-lysine and N delta-(2,3-diaminopropionyl)-D-ornithine were used as carriers for one or two cytotoxic moieties. The enhanced biological activities produced by the incorporation of D amino acids into position 6 of the agonistic analogues were further increased by the attachment of hydrophobic cytotoxic groups, resulting in compounds with 10-50 times higher activity than LH-RH. Most of the monosubstituted agonistic analogues showed high affinities for the membrane receptors of human breast cancer cells, while the receptor binding affinities of peptides containing two cytotoxic side chains were lower. Antagonistic carriers [Ac-D-Nal(2)1,D-Phe(4Cl)2,D-Trp3,Arg5,D-Lys6,D-Ala10] LH-RH [where Nal(2) is 3-(2-naphthyl)alanine], [Ac-D-Nal(2)1,D-Phe(4Cl)2,D-Trp3,Arg5,N epsilon-(2,3-diaminopropionyl)-D-Lys6,D-Ala10]LH-RH, and their D-Pal(3)3 homologs [Pal(3) is 3-(3-pyridyl)alanine] as well as [Ac-D-Nal(2)1,D-Phe(4Cl)2,D-Pal(3)3,Tyr5,N epsilon-(2,3-diamino-propionyl)-D-Lys6,D-Ala10]LH-RH were linked to cytotoxic compounds. The hybrid molecules inhibited ovulation in rats at doses of 10 micrograms and suppressed LH release in vitro. The receptor binding of cytotoxic analogues was decreased compared to the precursor peptides, although analogues with 2-(hydroxymethyl)anthraquinone hemiglutarate had high affinities. All of the cytotoxic analogues tested inhibited [3H]thymidine incorporation into DNA in cultures of human breast and prostate cancer cell lines

  16. Luteinizing hormone (LH)-responsive Cushing's syndrome: the demonstration of LH receptor messenger ribonucleic acid in hyperplastic adrenal cells, which respond to chorionic gonadotropin and serotonin agonists in vitro

    NARCIS (Netherlands)

    R.A. Feelders (Richard); W.W. de Herder (Wouter); S.W.J. Lamberts (Steven); L.J. Hofland (Leo); P.M. van Koetsveld (Peter); M. Verhoef-Post (Miriam); A.P.N. Themmen (Axel); F.H. de Jong (Frank); H.J. Bonjer (Jaap); A.J. Clark (Adrian); A-J. van der Lely (Aart-Jan)

    2003-01-01

    textabstractIn a substantial part of adrenal adenomas and hyperplasias from patients with Cushing's syndrome, cortisol production is controlled by the expression of aberrant hormone receptors on adrenocortical cells. We present in vivo and in vitro data of two patients with a

  17. Internalization of Rat FSH and LH/CG Receptors by rec-eCG in CHO-K1 Cells.

    Science.gov (United States)

    Park, Jong-Ju; Seong, Hun-Ki; Kim, Jeong-Soo; Munkhzaya, Byambaragchaa; Kang, Myung-Hwa; Min, Kwan-Sik

    2017-06-01

    Equine chorionic gonadotropin (eCG) is a unique molecule that elicits the response characteristics of both follicle-stimulating hormone (FSH) and luteinizing hormone (LH) in other species. Previous studies from this laboratory had demonstrated that recombinant eCG (rec-eCG) from Chinese hamster ovary (CHO-K1) cells exhibited both FSH- and LH-like activity in rat granulosa and Leydig cells. In this study, we analyzed receptor internalization through rec-eCGs, wild type eCG (eCGβ/α) and mutant eCG (eCGβ/αΔ56) with an N-linked oligosaccharide at Asn 56 of the α-subunit. Both the rec-eCGs were obtained from CHO-K1 cells. The agonist activation of receptors was analyzed by measuring stimulation time and concentrations of rec-eCGs. Internalization values in the stably selected rat follicle-stimulating hormone receptor (rFSHR) and rat luteinizing/chorionic gonadotropin receptor (rLH/CGR) were highest at 50 min after stimulation with 10 ng of rec-eCGβ/α. The dose-dependent response was highest when 10 ng of rec-eCGβ/α was used. The deglycosylated eCGβ/αΔ56 mutant did not enhance the agonist-stimulated internalization. We concluded that the state of activation of rFSHR and rLH/CGR could be modulated through agonist-stimulated internalization. Our results suggested that the eLH/CGRs are mostly internalized within 60 min by agonist-stimulation by rec-eCG. We also suggested that the lack of responsiveness of the deglycosylated eCGβ/ αΔ56 was likely because the site of glycosylation played a pivotal role in agonist-stimulated internalization in cells expressing rFSHR and rLH/CGR.

  18. 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.

  19. The Multiple Facets of Lutein: A Call for Further Investigation in the Perinatal Period

    OpenAIRE

    Perrone, Serafina; Tei, Monica; Longini, Mariangela; Buonocore, Giuseppe

    2016-01-01

    Lutein may have important antioxidant actions in free-radical-mediated diseases, in addition to its well-known antioxidant and cytoprotective effects on macula and photoreceptors. The peculiar perinatal susceptibility to oxidative stress indicates that prophylactic use of antioxidants as lutein could help to prevent or at least to reduce oxidative stress related diseases in newborns. Since lutein is not synthesized by humans, the intake primarily depends on diet or supplementation. Newborns r...

  20. The effect of Mono- and Diglycerides on the Digestion and Absorption of Lutein in Lymph Fistula Rats.

    Science.gov (United States)

    Tso, Patrick; Vurma, Mustafa; Ko, Chih-Wei; Lee, Dana M; DeMichele, Stephen

    2018-02-22

    Breastmilk lutein is better absorbed by infants than lutein delivered in infant formula. Therefore, we wish to better understand the possible absorption differences of lutein in breast milk versus infant formula by determining its bioavailability after gastric administration and whether the intestinal absorption of lutein can be improved by using new delivery vehicles. STUDY ONE compared the intestinal uptake, and the lymphatic and portal transport of lutein in conscious lymph fistula rats. Four groups of lymph and portal vein cannulated rats (n = 8-10/group) were randomized to receive via the gastric tube increasing doses (10,20,40,or 80mg/kg) of 20% lutein in safflower oil (SO) suspension to assess whether there was a saturable level of lutein that can be absorbed and transported in lymph. Aliquots of hourly portal blood and lymph were taken for lutein analysis. The dose response study showed that 20 mg/kg lutein was the saturable level of lymphatic lutein absorption with no lutein detected in portal circulation at any dosage level tested. STUDY TWO randomized five groups of lymph fistula rats (n = 4-9/group) to receive 20 mg/kg lutein from either lutein in SO or from lutein in four different mono- and diglyceride oil mixtures (MDG). Gastric infusion of lutein suspended in MDG (20 mg/kg) significantly improved (71% - 211%; plutein output 2-6 hours after lipid feeding versus lutein in SO. We conclude that a mixture of MDG helps solubilize lutein and facilitate gastrointestinal micelle formation thus improving lymphatic lutein absorption compared to triglyceride oils.

  1. Bioconversion of lutein by Enterobacter hormaechei to form a new compound, 8-methyl-α-ionone.

    Science.gov (United States)

    Zhong, Guifang; Wang, Fangfang; Sun, Jianhong; Ye, Jianbin; Mao, Duobin; Ma, Ke; Yang, Xuepeng

    2017-07-01

    To investigate the final product of the bioconversion of lutein by a novel lutein-degrading bacterium, Enterobacter hormaechei A20, and the kinetics of the process. A new product, 8-methyl-α-ionone, was resolved by GC-MS. The compound was further identified by NMR. A conversion yield of 90% was achieved by E. hormaechei in 36 h with 10 g lutein l -1 . This is the first report of the bioconversion of lutein to form 8-methyl-α-ionone. A degradation pathway is proposed.

  2. Transference of lutein during cheese making, color stability, and sensory acceptance of prato cheese

    Directory of Open Access Journals (Sweden)

    Mirian Tiaki Kaneiwa Kubo

    2013-02-01

    Full Text Available The consumption of lutein is associated with the prevention and reduction of age-related macular degeneration. Its incorporation into Prato cheese as a yellowish food coloring is a valid alternative to increase the daily intake of this compound. However, part of the lutein added may be lost in the whey during the cheese making, or it can be degraded by light during storage, resulting in color changes reducing the sensory acceptance of the cheese. The objectives of this study were to determine the transference of the lutein (dye, added to the milk, in the whey, and cheese, to evaluate the effect of the lutein addition, light exposure, and storage time on the cheese color, and to verify the sensory acceptance of Prato cheese with addition of lutein. The lutein recovery of cheese was 95.25%. Color saturation (chrome increased during storage time resulting in a cheese with more intense color, but there were no changes in the hue of the cheese. Adjusting the amount of lutein added to Prato cheese may lead to greater acceptance. The high recovery of lutein in the cheese and the fact that the hue remained unchanged during storage under light showed that the incorporation of lutein into Prato cheese is feasible from a technical point of view.

  3. Dietary effects of lutein-fortified chlorella on milk components of Holstein cows.

    Science.gov (United States)

    Jeon, Jin-Young; Park, Keun-Kyu; Lee, Kyung-Woo; Jang, Seung-Wan; Moon, Byung-Hern; An, Byoung-Ki

    2016-01-01

    This study was conducted to investigate the dietary effect of conventional or lutein-fortified chlorella on milk production and lutein incorporation in milk. Fifteen Holstein cows in mid-lactation were used in a 3 × 3 Latin square design each with a 21-day period. Cows were top-dressed daily with 30 g of conventional or lutein-fortified chlorella for 3 weeks. Cows without chlorella served as the control. The feed intake and milk yield were not affected by dietary treatments. The concentrations of milk protein and solids non-fat in groups fed diets containing both conventional and lutein-fortified chlorella were significantly higher than those of the control group (P milk fat among groups. The levels of plasma glutamic oxaloacetic transaminase, glutamic pyruvic transaminase, interferon-gamma and interleukin-2 were not influenced by the dietary treatments. Lutein content in milk was significantly increased in groups fed lutein-fortified chlorella as compared with those of conventional chlorella and control, respectively (P lutein-fortified chlorella has positive effects on milk components and the use of lutein-fortified chlorella in a dairy diet is effective in the production of milk enriched with lutein.

  4. A regulator of G Protein signaling, RGS3, inhibits gonadotropin-releasing hormone (GnRH-stimulated luteinizing hormone (LH secretion

    Directory of Open Access Journals (Sweden)

    Musgrove Lois C

    2001-11-01

    Full Text Available Abstract Background Luteinizing hormone secreted by the anterior pituitary gland regulates gonadal function. Luteinizing hormone secretion is regulated both by alterations in gonadotrope responsiveness to hypothalamic gonadotropin releasing hormone and by alterations in gonadotropin releasing hormone secretion. The mechanisms that determine gonadotrope responsiveness are unknown but may involve regulators of G protein signaling (RGSs. These proteins act by antagonizing or abbreviating interaction of Gα proteins with effectors such as phospholipase Cβ. Previously, we reported that gonadotropin releasing hormone-stimulated second messenger inositol trisphosphate production was inhibited when RGS3 and gonadotropin releasing hormone receptor cDNAs were co-transfected into the COS cell line. Here, we present evidence for RGS3 inhibition of gonadotropin releasing hormone-induced luteinizing hormone secretion from cultured rat pituitary cells. Results A truncated version of RGS3 (RGS3T = RGS3 314–519 inhibited gonadotropin releasing hormone-stimulated inositol trisphosphate production more potently than did RSG3 in gonadotropin releasing hormone receptor-bearing COS cells. An RSG3/glutathione-S-transferase fusion protein bound more 35S-Gqα than any other member of the G protein family tested. Adenoviral-mediated RGS3 gene transfer in pituitary gonadotropes inhibited gonadotropin releasing hormone-stimulated luteinizing hormone secretion in a dose-related fashion. Adeno-RGS3 also inhibited gonadotropin releasing hormone stimulated 3H-inositol phosphate accumulation, consistent with a molecular site of action at the Gqα protein. Conclusions RGS3 inhibits gonadotropin releasing hormone-stimulated second messenger production (inositol trisphosphate as well as luteinizing hormone secretion from rat pituitary gonadotropes apparently by binding and suppressing the transduction properties of Gqα protein function. A version of RGS3 that is amino

  5. Differentiation between lutein monoester regioisomers and detection of lutein diesters from marigold flowers (Tagetes erecta L.) and several fruits by liquid chromatography-mass spectrometry.

    Science.gov (United States)

    Breithaupt, Dietmar E; Wirt, Ursula; Bamedi, Ameneh

    2002-01-02

    Liquid chromatography-atmospheric pressure chemical ionization mass spectrometry (LC-APCIMS) was employed for the identification of eight lutein monoesters, formed by incomplete enzymatic saponification of lutein diesters of marigold (Tagetes erecta L.) by Candida rugosa lipase. Additionally, the main lutein diesters naturally occurring in marigold oleoresin were chromatographically separated and identified. The LC-MS method allows for characterization of lutein diesters occurring as minor components in several fruits; this was demonstrated by analysis of extracts of cape gooseberry (Physalis peruviana L.), kiwano (Cucumis metuliferus E. Mey. ex Naud.), and pumpkin (Cucurbita pepo L.). The assignment of the regioisomers of lutein monoesters is based on the characteristic fragmentation pattern: the most intense daughter ion generally results from the loss of the substituent (fatty acid or hydroxyl group) bound to the epsilon-ionone ring, yielding an allylic cation. The limit of detection was estimated at 0.5 microg/mL with lutein dimyristate as reference compound. This method provides a useful tool to obtain further insight into the biochemical reactions leading to lutein ester formation in plants.

  6. 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.

  7. Radioimmunological and clinical studies with luteinizing hormone releasing hormone (LRH)

    International Nuclear Information System (INIS)

    Dahlen, H.G.

    1986-01-01

    Radioimmunoassay for Luteinizing Hormone Releasing Hormone (LRH) has been established, tested and applied. Optimal conditions for the performance with regards to incubation time, incubation temperature, concentration of antiserum and radiolabelled LRH have been established. The specificity of the LRH immunoassay was investigated. Problems with direct measurement of LRH in plasmas of radioimmunoassay are encountered. The LRH distribution in various tissues of the rat are investigated. By means of a system for continuous monitoring of LH and FSH in women the lowest effective dose of LRH causing a significant release of LH and FSH could be established. (Auth.)

  8. Conservative Management of Theca Lutein Cyst Accident: A Case Report

    Directory of Open Access Journals (Sweden)

    Radha Bai Prabhu T

    2017-10-01

    Full Text Available Theca lutein cysts can occur in 20-25% of molar pregnancies. These cysts can undergo complications such as torsion, rupture, and haemorrhage. As these are functional cysts, when there are complications such as torsion they can be managed conservatively by aspirating the cysts under ultrasound guidance or by detorsion at the time of laparoscopy. By simple detorsion, ovaries can be preserved in 80-90% of cases. In order to prevent recurrence adnexal fixation can be undertaken by plicating the ovarian ligament.

  9. Ultrastructural and biochemical evidence for the presence of mature steroidogenic acute regulatory protein (StAR) in the cytoplasm of human luteal cells.

    Science.gov (United States)

    Sierralta, Walter D; Kohen, Paulina; Castro, Olga; Muñoz, Alex; Strauss, Jerome F; Devoto, Luigi

    2005-10-20

    The distribution of the steroidogenic acute regulatory protein (StAR) inside thecal and granulosa-lutein cells of human corpus luteum (CL) was assessed by immunoelectron microscopy. We found greater levels of StAR immunolabeling in steroidogenic cells from early- and mid-than in late luteal phase CL and lower levels in cells from women treated with a GnRH antagonist in the mid-luteal phase. Immunoelectron microscopy revealed significant levels of StAR antigen in the mitochondria and in the cytoplasm of luteal cells. The 30 kDa mature StAR protein was present in both mitochondria and cytosol (post-mitochondrial) fractions from homogenates of CL at different ages, whereas cytochrome c and mitochondrial HSP70 were detected only in the mitochondrial fraction. Therefore, we hypothesized that either appreciable processing of StAR 37 kDa pre-protein occurs outside the mitochondria, or mature StAR protein is selectively released into the cytoplasm after mitochondrial processing. The presence of mature StAR in the cytoplasm is consonant with the notion that StAR acts on the outer mitochondrial membrane to effect sterol import, and that StAR may interact with other cytoplasmic proteins involved in cholesterol metabolism, including hormone sensitive lipase.

  10. 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.

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

    NARCIS (Netherlands)

    Miranda, B.E.C.; 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

  12. 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.

  13. 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...

  14. Long-term oral feeding of lutein-fortified milk increases voluntary running distance in rats.

    Directory of Open Access Journals (Sweden)

    Megumi Matsumoto

    Full Text Available To evaluate the effects of lutein-fortified milk administration on running exercise, a voluntary wheel-running model was performed in rats. Four-week-old F344 rats were administered test milk (10 mL/kg daily following a 4-h fasting period, and their running distances were measured each day for a 9-week period. Total weekly running distance significantly increased from the sixth week until the end of the test period in lutein-supplemented rats (lutein-fortified milk administered compared with control rats (vehicle administered. This increase was not apparent in rats administered lutein alone. In the lutein-fortified-milk exercise group compared with the sedentary control group, carnitine palitroyltransferase 1 (CPT-1, total AMP-activated protein kinase (tAMPK, and phosphorylated AMP-activated protein kinase (pAMPK contents were significantly increased in the gastrocnemius muscle, with a concomitant decrease in triglyceride and total cholesterol levels in the blood and liver. Furthermore, the lutein level in blood of lutein-administered rats significantly decreased with exercise. These results suggest that lutein-fortified milk may enhance the effect of exercise by effective utilization of lipids when combined with voluntary running.

  15. Long-term oral feeding of lutein-fortified milk increases voluntary running distance in rats.

    Science.gov (United States)

    Matsumoto, Megumi; Hagio, Masahito; Inoue, Ryo; Mitani, Tomohiro; Yajima, Masako; Hara, Hiroshi; Yajima, Takaji

    2014-01-01

    To evaluate the effects of lutein-fortified milk administration on running exercise, a voluntary wheel-running model was performed in rats. Four-week-old F344 rats were administered test milk (10 mL/kg) daily following a 4-h fasting period, and their running distances were measured each day for a 9-week period. Total weekly running distance significantly increased from the sixth week until the end of the test period in lutein-supplemented rats (lutein-fortified milk administered) compared with control rats (vehicle administered). This increase was not apparent in rats administered lutein alone. In the lutein-fortified-milk exercise group compared with the sedentary control group, carnitine palitroyltransferase 1 (CPT-1), total AMP-activated protein kinase (tAMPK), and phosphorylated AMP-activated protein kinase (pAMPK) contents were significantly increased in the gastrocnemius muscle, with a concomitant decrease in triglyceride and total cholesterol levels in the blood and liver. Furthermore, the lutein level in blood of lutein-administered rats significantly decreased with exercise. These results suggest that lutein-fortified milk may enhance the effect of exercise by effective utilization of lipids when combined with voluntary running.

  16. The Multiple Facets of Lutein: A Call for Further Investigation in the Perinatal Period.

    Science.gov (United States)

    Perrone, Serafina; Tei, Monica; Longini, Mariangela; Buonocore, Giuseppe

    Lutein may have important antioxidant actions in free-radical-mediated diseases, in addition to its well-known antioxidant and cytoprotective effects on macula and photoreceptors. The peculiar perinatal susceptibility to oxidative stress indicates that prophylactic use of antioxidants as lutein could help to prevent or at least to reduce oxidative stress related diseases in newborns. Since lutein is not synthesized by humans, the intake primarily depends on diet or supplementation. Newborns receive lutein exclusively from breast milk. Lutein supplementation in term newborns has been reported to reduce oxidative stress and increase antioxidant capacities in the first days of life. Innovative frontiers concerning lutein supplementation are orientated toward cardiometabolic health improvement and cognitive benefits. The safety of lutein as an antioxidant agent has been confirmed in experimental and clinical studies, but its routine use is not recommended in perinatal period. This review summarizes what is known about the role of lutein as an antioxidant and anti-inflammatory agent in animal model and humans.

  17. The Multiple Facets of Lutein: A Call for Further Investigation in the Perinatal Period

    Directory of Open Access Journals (Sweden)

    Serafina Perrone

    2016-01-01

    Full Text Available Lutein may have important antioxidant actions in free-radical-mediated diseases, in addition to its well-known antioxidant and cytoprotective effects on macula and photoreceptors. The peculiar perinatal susceptibility to oxidative stress indicates that prophylactic use of antioxidants as lutein could help to prevent or at least to reduce oxidative stress related diseases in newborns. Since lutein is not synthesized by humans, the intake primarily depends on diet or supplementation. Newborns receive lutein exclusively from breast milk. Lutein supplementation in term newborns has been reported to reduce oxidative stress and increase antioxidant capacities in the first days of life. Innovative frontiers concerning lutein supplementation are orientated toward cardiometabolic health improvement and cognitive benefits. The safety of lutein as an antioxidant agent has been confirmed in experimental and clinical studies, but its routine use is not recommended in perinatal period. This review summarizes what is known about the role of lutein as an antioxidant and anti-inflammatory agent in animal model and humans.

  18. Lutein Is Differentially Deposited across Brain Regions following Formula or Breast Feeding of Infant Rhesus Macaques.

    Science.gov (United States)

    Jeon, Sookyoung; Ranard, Katherine M; Neuringer, Martha; Johnson, Emily E; Renner, Lauren; Kuchan, Matthew J; Pereira, Suzette L; Johnson, Elizabeth J; Erdman, John W

    2018-01-01

    Lutein, a yellow xanthophyll, selectively accumulates in primate retina and brain. Lutein may play a critical role in neural and retinal development, but few studies have investigated the impact of dietary source on its bioaccumulation in infants. We explored the bioaccumulation of lutein in infant rhesus macaques following breastfeeding or formula-feeding. From birth to 6 mo of age, male and female rhesus macaques (Macaca mulatta) were either breastfed (BF) (n = 8), fed a formula supplemented with lutein, zeaxanthin, β-carotene, and lycopene (237, 19.0, 74.2, and 338 nmol/kg, supplemented formula-fed; SF) (n = 8), or fed a formula with low amounts of these carotenoids (38.6, 2.3, 21.5, and 0 nmol/kg, unsupplemented formula-fed; UF) (n = 7). The concentrations of carotenoids in serum and tissues were analyzed by HPLC. At 6 mo of age, the BF group exhibited significantly higher lutein concentrations in serum, all brain regions, macular and peripheral retina, adipose tissue, liver, and other tissues compared to both formula-fed groups (P Lutein concentrations were higher in the SF group than in the UF group in serum and all tissues, with the exception of macular retina. Lutein was differentially distributed across brain areas, with the highest concentrations in the occipital cortex, regardless of the diet. Zeaxanthin was present in all brain regions but only in the BF infants; it was present in both retinal regions in all groups but was significantly enhanced in BF infants compared to either formula group (P lutein concentrations compared to unsupplemented formula, concentrations were still well below those in BF infants. Regardless of diet, occipital cortex showed selectively higher lutein deposition than other brain regions, suggesting lutein's role in visual processing in early life. © 2018 American Society for Nutrition. All rights reserved.

  19. 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...

  20. Short-chain analogs of luteinizing hormone-releasing hormone containing cytotoxic moieties.

    Science.gov (United States)

    Janáky, T; Juhász, A; Rékási, Z; Serfözö, P; Pinski, J; Bokser, L; Srkalovic, G; Milovanovic, S; Redding, T W; Halmos, G

    1992-11-01

    Five hexapeptide and heptapeptide analogs of luteinizing hormone-releasing hormone (LH-RH) were synthesized for use as carriers for cytotoxic compounds. These short analogs were expected to enhance target selectivity of the antineoplastic agents linked to them. Native LH-RH-(3-9) and LH-RH-(4-9) containing D-lysine and D-ornithine at position 6 were amidated with ethylamine and acylated on the N terminus. The receptor-binding affinity of one hexapeptide carrier AJ-41 (Ac-Ser-Tyr-D-Lys-Leu-Arg-Pro-NH-Et) to human breast cancer cell membranes was similar to that of [D-Trp6]LH-RH. Alkylating nitrogen mustards (melphalan, Ac-melphalan), anthraquinone derivatives including anticancer antibiotic doxorubicin, antimetabolite (methotrexate), and cisplatin-like platinum complex were linked to these peptides through their omega-amino group at position 6. The hybrid molecules showed no LH-RH agonistic activity in vitro and in vivo but had nontypical antagonistic effects on pituitary cells in vitro at the doses tested. These analogs showed a wide range of receptor-binding affinities to rat pituitaries and cell membranes of human breast cancer and rat Dunning prostate cancer. Several of these conjugates exerted some cytotoxic effects on MCF-7 breast cancer cell line.

  1. Lutein in food supplements available on the markets of the Viszegrad countries

    Directory of Open Access Journals (Sweden)

    Miroslav Šivel

    2014-11-01

    Full Text Available RP-HPLC method with UV-VIS detection was implemented for determination of contents of lutein in food supplements available on the markets in the Czech Republic, Slovakia, Poland, and Hungary. Altogether, 48 samples of food supplements in three dosage forms (22 samples of tablets, 18 samples of soft capsules, and 8 samples of hard capsules were analysed. The amounts of lutein specified by the producer complied with their real contents only in 7 samples of the food supplements. Lutein in soft capsules showed the highest stability against oxidation; lutein in tablets was more prone to oxidation and lutein in hard capsules was most susceptible to oxidation process. Out of 21 Czech products, only four fell into the category of satisfactory products, three of them were soft capsules and one was a tablet. Out of 27 products manufactured abroad, only three were evaluated as satisfactory products, all of them were soft capsules, out of 48 analysed food supplement samples just seven fell into the category of satisfactory preparations, eight were evaluated as less satisfactory preparations, five were found inadequate products and 28 samples were labelled unsatisfactory. Only one in six analyzed samples contained the amount of lutein specified by the manufacturer, almost 60% of monitored lutein containing food supplement samples fell into the unsatisfactory product category.

  2. Long-Term Oral Feeding of Lutein-Fortified Milk Increases Voluntary Running Distance in Rats

    OpenAIRE

    Matsumoto, Megumi; Hagio, Masahito; Inoue, Ryo; Mitani, Tomohiro; Yajima, Masako; Hara, Hiroshi; Yajima, Takaji

    2014-01-01

    To evaluate the effects of lutein-fortified milk administration on running exercise, a voluntary wheel-running model was performed in rats. Four-week-old F344 rats were administered test milk (10 mL/kg) daily following a 4-h fasting period, and their running distances were measured each day for a 9-week period. Total weekly running distance significantly increased from the sixth week until the end of the test period in lutein-supplemented rats (lutein-fortified milk administered) compared wit...

  3. Dietary effects of lutein-fortified chlorella on milk components of Holstein cows

    OpenAIRE

    Jeon, Jin-Young; Park, Keun-Kyu; Lee, Kyung-Woo; Jang, Seung-Wan; Moon, Byung-Hern; An, Byoung-Ki

    2016-01-01

    This study was conducted to investigate the dietary effect of conventional or lutein-fortified chlorella on milk production and lutein incorporation in milk. Fifteen Holstein cows in mid-lactation were used in a 3???3 Latin square design each with a 21-day period. Cows were top-dressed daily with 30?g of conventional or lutein-fortified chlorella for 3?weeks. Cows without chlorella served as the control. The feed intake and milk yield were not affected by dietary treatments. The concentration...

  4. Ontogenesis of neurons producing luteinizing hormone-releasing hormone (LHRH) in the nervus terminalis of the rat.

    Science.gov (United States)

    Schwanzel-Fukuda, M; Morrell, J I; Pfaff, D W

    1985-08-15

    Immunoreactive luteinizing hormone-releasing hormone (LHRH) was first detected at 15 days of gestation in ganglion cells associated with the peripheral, intracranial, and central parts of the nervus terminalis of the rat. LHRH was not detected in any other structure of the central nervous system at this age. In the 17-day-old fetal rat, 62% of the total LHRH-reactive neuronal population was found in ganglion cells of the nervus terminalis. At this same age, immunoreactive beta-luteinizing hormone (beta-LH) was first seen in gonadotropes of the anterior pituitary gland. At 19 days of gestation, 31% of the total number of LHRH-reactive neurons observed in the rat brain was found in the nervus terminalis, and immunoreactive processes were first seen in the organum vasculosum of the lamina terminalis and in the median eminence. Our data indicate that from 15 to 19 days of gestation the nervus terminalis is a principal source of LHRH in the fetal rat. Presence of the decapeptide in the nervus terminalis prior to appearance of beta-LH in the anterior pituitary suggests a possible role for LHRH in this system on maturation of the gonadotropes and differentiation of the brain-pituitary-gonadal axis.

  5. Exposure to α-Tocopherol, Lutein or Ascorbic Acid improve Cumulus Expansion, Viability and Maturation of Swine Oocytes

    Directory of Open Access Journals (Sweden)

    Ileana Miclea

    2010-05-01

    Full Text Available Protection of the fatty acid and lipid components of oocytes that render them susceptible to free radical or other oxidative injury may prevent the damage currently associated with culture. The goal of this study was to establish the influence of several α-tocopherol, lutein and ascorbic acid concentrations on swine oocyte maturation, viability and the function of cumulus cells in order to improve culture media. Pig oocytes were cultured for 45 hours at 37°C in 5% CO2 atmosphere; in M199 containing several α-tocopherol (5, 10, 20, 40, 80 μM, lutein (2.5, 4, 5, 8, 10 M or ascorbic acid (50, 150, 250, 500, 750 μM concentrations and cumulus expansion was assessed. Afterwards oocytes were coloured using FDA, PI and Hoechst 33258. The differences between treatments were analyzed by the analysis of variance and interpreted using the Newman-Keuls method. When cultured in α-tocopherol supplemented medium the number of expanded COCs to be scored as 3 was significantly greater (p<0.05 for the 5 and 40 μM concentrations. The addition of 8 M lutein to the maturation medium lead to a significant (p<0.05 increase in the number of COCs that were scored at 4. For both α-tocopherol and lutein additions the numbers of oocytes stained by FDA, as well as those stained by Hoechst were greater than the control without being statistically significant. When cultured in 150 and 500 μM ascorbic acid the percentages of COCs scored at 4 were significantly lower (p<0.05 than the control. Also, significantly (p<0.05 fewer oocytes were stained with FDA when matured in 500 μM. Differences between the control and the several concentrations were significant (p<0.05 for 150 and 750 μM and distinctly significant (p<0.01 for 250 μM.

  6. Synthesis and in vitro anti-cancer evaluation of luteinizing hormone-releasing hormone-conjugated peptide.

    Science.gov (United States)

    Deng, Xin; Qiu, Qianqian; Ma, Ke; Huang, Wenlong; Qian, Hai

    2015-11-01

    Luteinizing hormone-releasing hormone (LHRH) is a decapeptide hormone released from the hypothalamus and shows high affinity binding to the LHRH receptors. It is reported that several cancer cells also express LHRH receptors such as breast, ovarian, prostatic, bladder and others. In this study, we linked B1, an anti-cancer peptide, to LHRH and its analogs to improve the activity against cancer cells with LHRH receptor. Biological evaluation revealed that TB1, the peptide contains triptorelin sequence, present favorable anti-cancer activity as well as plasma stability. Further investigations disclosed that TB1 trigger apoptosis by activating the mitochondria-cytochrome c-caspase apoptotic pathway, it also exhibited the anti-migratory effect on cancer cells.

  7. 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.

  8. Regional differences in the pituitary distribution of luteinizing hormone in the gonadectomized and proestrous female rat

    Science.gov (United States)

    Previous data have shown regional differences in the presence of anterior pituitary luteinizing hormone (LH) that generally correlate with comparable disparities in the distribution of gonadotropes throughout the gland. In female rats, the differences are apparent over the estro...

  9. 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

  10. Transference of lutein during cheese making, color stability, and sensory acceptance of Prato cheese

    OpenAIRE

    Kubo, MTK; Maus, D; Xavier, AAO; Mercadante, AZ; Viotto, WH

    2013-01-01

    The consumption of lutein is associated with the prevention and reduction of age-related macular degeneration. Its incorporation into Prato cheese as a yellowish food coloring is a valid alternative to increase the daily intake of this compound. However, part of the lutein added may be lost in the whey during the cheese making, or it can be degraded by light during storage, resulting in color changes reducing the sensory acceptance of the cheese. The objectives of this study were to determine...

  11. Why has Nature Chosen Lutein and Zeaxanthin to Protect the Retina?

    OpenAIRE

    Widomska, Justyna; Subczynski, Witold K

    2014-01-01

    Age-related macular degeneration (AMD) is associated with a low level of macular carotenoids in the eye retina. Only two carotenoids, namely lutein and zeaxanthin are selectively accumulated in the human eye retina from blood plasma where more than twenty other carotenoids are available. The third carotenoid which is found in the human retina, meso-zeaxanthin is formed directly in the retina from lutein. All these carotenoids, named also macular xanthophylls, play key roles in eye health and ...

  12. Lutein-fortified infant formula fed to healthy term infants: evaluation of growth effects and safety

    Directory of Open Access Journals (Sweden)

    Davis Anne M

    2010-05-01

    Full Text Available Abstract Background/Objectives Breast milk contains lutein derived from the mother's diet. This carotenoid is currently not added to infant formula, which has a small and variable lutein content from innate ingredients. This study was conducted to compare the growth of infants fed lutein-fortified infant formula with that of infants fed infant formula without lutein fortification. Subjects/Methods This 16-week study was prospective, randomized, controlled, and double-blind with parallel groups of healthy term infants fed either control formula (Wyeth S-26 Gold, designated as Gold or experimental formula (Wyeth S-26 Gold fortified with lutein at 200 mcg/l, designated as Gold + Lutein. Two hundred thirty-two (232 infants ≤ 14 days postnatal age were randomized and 220 (94.8% completed the study. Weight (g, head circumference (cm, and length (cm were measured at Weeks 4, 8, 12, and 16. The primary endpoint was weight gain (g/day from baseline to Week 16. Safety was assessed through monitoring of study events (SEs throughout the study and evaluation of selected blood chemistry tests performed at Week 16. Results Infants in both treatment groups demonstrated appropriate growth. No differences between treatment groups were found in any of the measures of growth at any of the measurement time points. Both study formulas were well tolerated. The mean values of all measured blood chemistry parameters fell within the modified normal ranges for infants, and the values for both groups for any measured parameter were similar. Conclusions Infants fed lutein-fortified S-26 Gold demonstrated growth equivalent to that of infants fed unfortified lutein formula.

  13. Synthesis and release of luteinizing hormone in vitro: manipulations of Ca2+ environment

    International Nuclear Information System (INIS)

    Liu, T.C.; Jackson, G.L.

    1985-01-01

    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 [ 3 H]glucosamine (glycosylation) and [ 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 [ 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

  14. Occurrence of neoxanthin and lutein epoxide cycle in parasitic Cuscuta species.

    Science.gov (United States)

    Kruk, Jerzy; Szymańska, Renata

    2008-01-01

    In the present study, xanthophyll composition of eight parasitic Cuscuta species under different light conditions was investigated. Neoxanthin was not detected in four of the eight species examined, while in others it occurred at the level of several percent of total xanthophylls. In C. gronovii and C. lupuliformis it was additionally found that the neoxanthin content was considerably stimulated by strong light. In dark-adapted plants, lutein epoxide level amounted to 10-22% of total xanthophylls in only three species, the highest being for C. lupuliformis, while in others it was below 3%, indicating that the lutein epoxide cycle is limited to only certain Cuscuta species. The obtained data also indicate that the presence of the lutein epoxide cycle and of neoxanthin is independent and variable among the Cuscuta species. The xanthophyll cycle carotenoids violaxanthin, antheraxanthin and zeaxanthin were identified in all the examined species and occurred at the level found in other higher plants. The xanthophyll and lutein epoxide cycle pigments showed typical response to high light stress. The obtained results also suggest that the ability of higher plants to synthesize lutein epoxide probably does not depend on the substrate specificity of zeaxanthin epoxidase but on the availability of lutein for the enzyme.

  15. Impact of Lutein Intervention in Mice on the Radiation Induced Clastogenic Changes

    Directory of Open Access Journals (Sweden)

    Vidya Vasudeva

    2017-10-01

    Full Text Available One of the genetic effects of radiation is that it may lead to formation of single or double strand breaks in DNA which can be observed in differentially stained polychromatic or normochromatic erythrocytes (PCE and NCE respectively. In pursuit of finding a natural radioprotector to treat the radiation induced damages; lutein, a carotenoid pigment is one such approach. Swiss albino mice are administered with the compound (lutein/gallic acid/DMSO with respective controls for 15 consecutive days after which they are irradiated. The whole blood is drawn for comet assay and the femur of the leg is removed to flush out the content of the bone marrow in BSA for the micronucleus assay. The comet slides are observed under the fluorescent microscope and the PCE/NCE or micronucleated PCEs or NCEs are scored blindly. Lutein in the present study has effectively reduced the olive moment and the tail moment. However, % DNA in tail has been maintained to normal levels in comparison to its control indicating lesser extent of damage to the genetic material. The percent micronucleated NCE (MnNCE has been decreased in the group treated with lutein prior to radiation. The % MnPCE and the PCE/(PCE + NCE ratio has been increased in all the irradiated groups; however lutein treatment has not drastically increased the formation of micronuclei in comparison to its control. This indicates that lutein shows a protective effect against the radiation induced cytogenetic damages in Swiss albino mice.

  16. Lutein Esterification in Wheat Flour Increases the Carotenoid Retention and Is Induced by Storage Temperatures

    Directory of Open Access Journals (Sweden)

    Elena Mellado-Ortega

    2017-12-01

    Full Text Available The present study aimed to evaluate the effects of long-term storage on the carotenoid pigments present in whole-grain flours prepared from durum wheat and tritordeum. As expected, higher storage temperatures showed a catabolic effect, which was very marked for free carotenoid pigments. Surprisingly, for both cereal genotypes, the thermal conditions favoured the synthesis of lutein esters, leading to an enhanced stability, slower degradation, and, subsequently, a greater carotenoid retention. The putative involvement of lipase enzymes in lutein esterification in flours is discussed, particularly regarding the preferential esterification of the hydroxyl group with linoleic acid at the 3′ in the ε-ring of the lutein molecule. The negative effects of processing on carotenoid retention were less pronounced in durum wheat flours, which could be due to an increased esterifying activity (the de novo formation of diesterified xanthophylls was observed. Moreover, clear differences were observed for tritordeum depending on whether the lutein was in a free or esterified state. For instance, lutein-3′-O-monolinoleate showed a three-fold lower degradation rate than free lutein at 37 °C. In view of our results, we advise that the biofortification research aimed at increasing the carotenoid contents in cereals should be based on the selection of varieties with an enhanced content of esterified xanthophylls.

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

    Directory of Open Access Journals (Sweden)

    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.

  18. Germ cells are not required to establish the female pathway in mouse fetal gonads.

    Directory of Open Access Journals (Sweden)

    Danielle M Maatouk

    Full Text Available The fetal gonad is composed of a mixture of somatic cell lineages and germ cells. The fate of the gonad, male or female, is determined by a population of somatic cells that differentiate into Sertoli or granulosa cells and direct testis or ovary development. It is well established that germ cells are not required for the establishment or maintenance of Sertoli cells or testis cords in the male gonad. However, in the agametic ovary, follicles do not form suggesting that germ cells may influence granulosa cell development. Prior investigations of ovaries in which pre-meiotic germ cells were ablated during fetal life reported no histological changes during stages prior to birth. However, whether granulosa cells underwent normal molecular differentiation was not investigated. In cases where germ cell loss occurred secondary to other mutations, transdifferentiation of granulosa cells towards a Sertoli cell fate was observed, raising questions about whether germ cells play an active role in establishing or maintaining the fate of granulosa cells. We developed a group of molecular markers associated with ovarian development, and show here that the loss of pre-meiotic germ cells does not disrupt the somatic ovarian differentiation program during fetal life, or cause transdifferentiation as defined by expression of Sertoli markers. Since we do not find defects in the ovarian somatic program, the subsequent failure to form follicles at perinatal stages is likely attributable to the absence of germ cells rather than to defects in the somatic cells.

  19. Germ Cells Are Not Required to Establish the Female Pathway in Mouse Fetal Gonads

    Science.gov (United States)

    Maatouk, Danielle M.; Mork, Lindsey; Hinson, Ashley; Kobayashi, Akio; McMahon, Andrew P.; Capel, Blanche

    2012-01-01

    The fetal gonad is composed of a mixture of somatic cell lineages and germ cells. The fate of the gonad, male or female, is determined by a population of somatic cells that differentiate into Sertoli or granulosa cells and direct testis or ovary development. It is well established that germ cells are not required for the establishment or maintenance of Sertoli cells or testis cords in the male gonad. However, in the agametic ovary, follicles do not form suggesting that germ cells may influence granulosa cell development. Prior investigations of ovaries in which pre-meiotic germ cells were ablated during fetal life reported no histological changes during stages prior to birth. However, whether granulosa cells underwent normal molecular differentiation was not investigated. In cases where germ cell loss occurred secondary to other mutations, transdifferentiation of granulosa cells towards a Sertoli cell fate was observed, raising questions about whether germ cells play an active role in establishing or maintaining the fate of granulosa cells. We developed a group of molecular markers associated with ovarian development, and show here that the loss of pre-meiotic germ cells does not disrupt the somatic ovarian differentiation program during fetal life, or cause transdifferentiation as defined by expression of Sertoli markers. Since we do not find defects in the ovarian somatic program, the subsequent failure to form follicles at perinatal stages is likely attributable to the absence of germ cells rather than to defects in the somatic cells. PMID:23091613

  20. Mioblastoma de células granulosas de la base de la lengua en el recién nacido

    Directory of Open Access Journals (Sweden)

    Manuel Estrada Sarmiento

    2002-09-01

    Full Text Available Se realiza una revisión bibliográfica del mioblastoma de células granulosas de la lengua, sus características clínicas y diagnóstico diferencial. Se hace énfasis en la posibilidad real de su presentación frente a un síndrome de dificultad respiratoria del recién nacido. Se presenta a un enfermo con estas características, el cual fue intervenido quirúrgicamente por un grupo multidisciplinario en nuestra institución. El paciente tuvo una evolución posoperatoria libre de complicaciones y se le dio de alta curado.A bibiliographic review of the granular cell tumor of the tongue, its clinical characteristics and differential diagnosis is made. The real possibility of its appearance in the face of a respiratory distress syndrome in a newborn infant is emphasized. A patient with these characteristics that was operated on by a multidisciplinary team in our institution is presented. The patient had a postoperative evolution with no complications and he was discharged completely cured.

  1. Possible influences of lutein and zeaxanthin on the developing retina.

    Science.gov (United States)

    Zimmer, J Paul; Hammond, Billy R

    2007-03-01

    The carotenoids lutein and zeaxanthin (LZ) are found throughout the central nervous system but reach their highest concentration within the macular region of the primate retina where they are commonly referred to as the macular pigments. Although LZ are a major integral feature of the central fovea, no information currently exists regarding the effects of variability in the concentration of these pigments on the developing retina. In particular, the long-term effects of very low levels of macular pigment are not known and potentially meaningful. Macular pigment levels depend upon dietary intake since LZ cannot be synthesized de novo. Infants with low intake of LZ (eg, infants receiving unfortified infant formula or breast milk from mothers with low carotenoid diets) would be expected to have considerably lower macular pigment compared with infants with high LZ intake (eg, breast-fed infants with mothers on carotenoid-rich diets). In this paper we discuss possible implications of this difference and the available evidence suggesting that LZ could influence the developing visual system.

  2. 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.

  3. Hubungan Kadar βHCG Praevakuasi, Gambaran Histopatologi, dan Kista Lutein dengan Performa βHCG pada Penderita Mola Hidatidosa yang Berkembang Menjadi PTG dan Kembali Normal

    Directory of Open Access Journals (Sweden)

    Yudi Mulyana Hidayat

    2014-12-01

    Full Text Available The incidence of trophoblastic diseases in Indonesia and developing countries is relatively high compared to the developed countries. The incidence of gestational trophoblast tumors (GTT after the evacuation of a hydatidiform mole ranges from 10% to 20%. Several clinical variables have been studied as the risk factors for malignancy, including the pre-evacuation level of beta human chorionic gonadotropin (βHCG, histopathological appearance, and the presence of lutein cysts. The purpose of this study was to determine the relationship between βHCG decline and pre-evacuation βHCG levels, histopathological features, and the lutein cysts status in patients with moles. This study was a case control study of patients with complete hydatidiform mole in Dr. Hasan Sadikin General Hospital during the period of 2007-2011. The results revealed that there was a significant correlation between the level of βHCG ≥100,000 mIU/mL and post-molar malignancy (p<0.05. There was also a significant relationship between the histopathologic feature of excessive post-molar cell proliferation and malignancy (p<0.05 and between the presence of lutein cyst and post-molar malignancy (p<0.05. This study concludes that the pre-evacuation βHCG level ≥100.000 mIU/mL, excessive proliferation, and the presence of lutein cysts are correlated with malignancy after molar evacuation. These risk factors are useful to differentiate whether a complete hydatidiform mole will become malignant or remain benign.

  4. Metabolism of Lutein and Zeaxanthin in Rhesus Monkeys: Identification of (3R,6′R)- and (3R,6′S)-3′-Dehydro-lutein as Common Metabolites and Comparison to Humans

    Science.gov (United States)

    Albert, Gesa I.; Hoeller, Ulrich; Schierle, Joseph; Neuringer, Martha; Johnson, Elizabeth J.; Schalch, Wolfgang

    2012-01-01

    Lutein and zeaxanthin are xanthophylls that can be found highly concentrated in the macula of the retina. They are thought to protect the macula through their role as blue-light filters and because of their antioxidant and singlet oxygen quenching properties. Examination of metabolites unique to lutein and zeaxanthin such as 3′-dehydro-lutein, and of their stereochemistry may provide insight to the mechanism by which they are formed and by which they exert protection. To evaluate the formation of such metabolites, eleven monkeys were raised on a xanthophyll-free diet, and supplemented with pure lutein or pure zeaxanthin (2.2 mg/kg body weight/d). The period of supplementation ranged between 12 to 92 weeks. At study start and throughout the study, serum samples were taken and analyzed for xanthophylls using different HPLC systems. Xanthophyll metabolites were identified using UV/VIS and HR-MS detection. Lutein and zeaxanthin metabolites were found in detectable amounts with 3′-dehydro-lutein being a common metabolite of both. Using chiral-phase HPLC, two diastereomers, (3R,6′R)-3′-dehydro-lutein and (3R,6′S)-3′-dehydro-lutein, were identified and shown to be present in nearly equimolar amounts. A pathway for their formation from either lutein or zeaxanthin is proposed. These finding were comparable to results obtained with human plasma. PMID:18582588

  5. Lutein accumulates in subcellular membranes of brain regions in adult rhesus macaques: Relationship to DHA oxidation products.

    Directory of Open Access Journals (Sweden)

    Emily S Mohn

    Full Text Available Lutein, a carotenoid with anti-oxidant functions, preferentially accumulates in primate brain and is positively related to cognition in humans. Docosahexaenoic acid (DHA, an omega-3 polyunsaturated fatty acid (PUFA, is also beneficial for cognition, but is susceptible to oxidation. The present study characterized the membrane distribution of lutein in brain regions important for different domains of cognitive function and determined whether membrane lutein was associated with brain PUFA oxidation.Adult rhesus monkeys were fed a stock diet (~2 mg/day lutein or ~0.5 μmol/kg body weight/day (n = 9 or the stock diet plus a daily supplement of lutein (~4.5 mg/day or~1 μmol/kg body weight/day and zeaxanthin (~0.5 mg/day or 0.1 μmol/kg body weight/day for 6-12 months (n = 4. Nuclear, myelin, mitochondrial, and neuronal plasma membranes were isolated using a Ficoll density gradient from prefrontal cortex (PFC, cerebellum (CER, striatum (ST, and hippocampus (HC. Carotenoids, PUFAs, and PUFA oxidation products were measured using HPLC, GC, and LC-GC/MS, respectively.All-trans-lutein (ng/mg protein was detected in all regions and membranes and was highly variable among monkeys. Lutein/zeaxanthin supplementation significantly increased total concentrations of lutein in serum, PFC and CER, as well as lutein in mitochondrial membranes and total DHA concentrations in PFC only (P<0.05. In PFC and ST, mitochondrial lutein was inversely related to DHA oxidation products, but not those from arachidonic acid (P <0.05.This study provides novel data on subcellular lutein accumulation and its relationship to DHA oxidation in primate brain. These findings support the hypothesis that lutein may be associated with antioxidant functions in the brain.

  6. Fat content affects bioaccessibility and efficiency of enzymatic hydrolysis of lutein esters added to milk and yogurt

    OpenAIRE

    Xavier, Ana Augusta Odorissi; Mercadante, Adriana Zerlotti; Garrido-Fernández, Juan; Pérez-Gálvez, Antonio

    2014-01-01

    © 2014 Elsevier Ltd. Addition of lutein to dairy products is an alternative that widens the range of foods which could be lutein sources. However, bioaccessibility is an essential aspect to be considered during the development of products with added bioactive substances. We evaluated the in vitro bioaccessibility of lutein esters added to milk and yogurt with different fat contents, and determined the efficiency of enzymatic hydrolysis of the esters during digestion. Bioaccessibility of lutei...

  7. 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.

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

    Science.gov (United States)

    Zamroziewicz, Marta K; Paul, Erick J; Zwilling, Chris E; Johnson, Elizabeth J; Kuchan, Matthew J; Cohen, Neal J; Barbey, Aron K

    2016-01-01

    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 76 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 and

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

    Directory of Open Access Journals (Sweden)

    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

  10. 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.

  11. The Adequate Corpus Luteum: miR-96 Promotes Luteal Cell Survival and Progesterone Production.

    Science.gov (United States)

    Mohammed, Bushra T; Sontakke, Sadanand D; Ioannidis, Jason; Duncan, W Colin; Donadeu, F Xavier

    2017-07-01

    Inadequate progesterone production from the corpus luteum is associated with pregnancy loss. Data available in model species suggest important roles of microRNAs (miRNAs) in luteal development and maintenance. To comprehensively investigate the involvement of miRNAs during the ovarian follicle-luteal transition. The effects of specific miRNAs on survival and steroid production by human luteinized granulosa cells (hLGCs) were tested using specific miRNA inhibitors. Candidate miRNAs were identified through microarray analyses of follicular and luteal tissues in a bovine model. An academic institution in the United Kingdom associated with a teaching hospital. hLGCs were obtained by standard transvaginal follicular-fluid aspiration from 35 women undergoing assisted conception. Inhibition of candidate miRNAs in vitro. Levels of miRNAs, mRNAs, FOXO1 protein, apoptosis, and steroids were measured in tissues and/or cultured cells. Two specific miRNA clusters, miR-183-96-182 and miR-212-132, were dramatically increased in luteal relative to follicular tissues. miR-96 and miR-132 were the most upregulated miRNAs within each cluster. Database analyses identified FOXO1 as a putative target of both these miRNAs. In cultured hLGCs, inhibition of miR-96 increased apoptosis and FOXO1 protein levels, and decreased progesterone production. These effects were prevented by small interfering RNA-mediated downregulation of FOXO1. In bovine luteal cells, miR-96 inhibition also led to increases in apoptosis and FOXO1 protein levels. miR-96 targets FOXO1 to regulate luteal development through effects on cell survival and steroid production. The miR-183-96-182 cluster could provide a novel target for the manipulation of luteal function. Copyright © 2017 Endocrine Society

  12. Obesity, lutein metabolism, and age-related macular degeneration: a web of connections.

    Science.gov (United States)

    Johnson, Elizabeth J

    2005-01-01

    Age-related macular degeneration (AMD) is a major cause of visual impairment in the United States. Currently there is no effective cure for this disease. Risk factors include decreased lutein and zeaxanthin status and obesity. Obesity is also an increasing public health concern. The alarming increase in the prevalence of obesity further exacerbates the public health concern of AMD. The mechanism by which obesity increases the risk of AMD may be related to the physiologic changes that occur with this condition. These include increased oxidative stress, changes in the lipoprotein profile, and increased inflammation. These changes would also result in an increased destruction and a decreased circulatory delivery of lutein and zeaxanthin to the macula of the eye. Therefore, the mechanism by which obesity is related to AMD risk may be through indirect effects on changes in lutein and zeaxanthin status and metabolism.

  13. Estradiol and luteinizing hormone regulate recognition memory following subchronic phencyclidine: Evidence for hippocampal GABA action.

    Science.gov (United States)

    Riordan, Alexander J; Schaler, Ari W; Fried, Jenny; Paine, Tracie A; Thornton, Janice E

    2018-05-01

    The cognitive symptoms of schizophrenia are poorly understood and difficult to treat. Estrogens may mitigate these symptoms via unknown mechanisms. To examine these mechanisms, we tested whether increasing estradiol (E) or decreasing luteinizing hormone (LH) could mitigate short-term episodic memory loss in a phencyclidine (PCP) model of schizophrenia. We then assessed whether changes in cortical or hippocampal GABA may underlie these effects. Female rats were ovariectomized and injected subchronically with PCP. To modulate E and LH, animals received estradiol capsules or Antide injections. Short-term episodic memory was assessed using the novel object recognition task (NORT). Brain expression of GAD67 was analyzed via western blot, and parvalbumin-containing cells were counted using immunohistochemistry. Some rats received hippocampal infusions of a GABA A agonist, GABA A antagonist, or GAD inhibitor before behavioral testing. We found that PCP reduced hippocampal GAD67 and abolished recognition memory. Antide restored hippocampal GAD67 and rescued recognition memory in PCP-treated animals. Estradiol prevented PCP's amnesic effect in NORT but failed to restore hippocampal GAD67. PCP did not cause significant differences in number of parvalbumin-expressing cells or cortical expression of GAD67. Hippocampal infusions of a GABA A agonist restored recognition memory in PCP-treated rats. Blocking hippocampal GAD or GABA A receptors in ovx animals reproduced recognition memory loss similar to PCP and inhibited estradiol's protection of recognition memory in PCP-treated animals. In summary, decreasing LH or increasing E can lessen short-term episodic memory loss, as measured by novel object recognition, in a PCP model of schizophrenia. Alterations in hippocampal GABA may contribute to both PCP's effects on recognition memory and the hormones' ability to prevent or reverse them. Copyright © 2018 Elsevier Ltd. All rights reserved.

  14. [Prevention and treatment of age-related macular degeneration by extract of Fructus lycii and its constituents lutein/zeaxanthin: an in vive and in vitro experimental research].

    Science.gov (United States)

    Huang, Bing-Lin; Ding, Shu-Hua; Hang, Li; Zheng, Shi-Zhong; Li, Wei; Xu, Xin-rong

    2013-04-01

    To investigate the in vivo inhibition of extract of Fructus lycii (FL) on the expressions of cathepsin B (Cat B) and cystatin C (Cys C) in high-fat diet and hydroquinone (HQ) induced model mice with age-related macular degeneration (AMD), and to explore the in vitro effects of lutein and zeaxanthin on hydrogen peroxide (H2O2,) induced expressions of matrix metalloproteinase 2 (MMP-2) and tissue inhibitor of metalloproteinase 2 (TIMP-2) on ARPE-19 cells. Fifty female 8-month-old C57BL/6 mice were recruited in this research. Ten mice fed with regular diet was taken as the age control group. The rest 40 mice were fed with high fat diet for 6 months, followed by adding HQ (0. 8%) in the drinking water for 3 consecutive months. Then the modeled mice were randomly divided into the model control group (n =10), the high (at the daily dose of 3.75 g/kg), middle (at the daily dose of 2.50 g/kg), and low dose (at the daily dose of 1.25 g/kg) FL groups, 10 in each group. The extract of FL at each dose was respectively administered to mice by gastrogavage for 3 successive months. By the end of the experiment, the mice were killed and their eyeballs were removed. The protein expressions of Cat B and Cys C were observed by immunohistochemical assay. The mRNA and protein expressions of Cat B and Cys C were detected by real-time PCR and Western blot respectively. The drug concentrations of H2O2, lutein, and zeaxanthin were screened and detected using the activity of cell proliferation. The protein expressions of MMP-2 and TIMP-2 were detected using Western blot. Compared with the age control group, the mRNA and protein expressions of Cat B and Cys C were significantly higher in the in vivo model control group (P <0.05, P <0.01). The mRNA expressions of Cat B and Cys C were weaker in the middle and high dose FL groups than in the model control group (P <0. 05, P <0. 01). In in vitro cells, lutein and zeaxanthin could down-regulate the protein expressions of MMP-2 and TIMP-2 in H202

  15. Delayed lactogenesis II secondary to gestational ovarian theca lutein cysts in two normal singleton pregnancies.

    Science.gov (United States)

    Hoover, Kathleen L; Barbalinardo, Laurie H; Platia, Maria Pia

    2002-08-01

    Hyperreactio luteinalis is an unusual condition in which, during pregnancy, both ovaries are enlarged by multiple theca lutein cysts that produce a high level of testosterone. Several weeks postpartum, the cysts resolve and testosterone level returns to normal. Two case studies are presented in which mothers with gestational ovarian theca lutein cysts experienced delayed lactogenesis II. The elevated testosterone at the time of birth suppressed milk production. Once the testosterone level dropped to approximately 300 ng/dL, milk production began. After the initial delay, both mothers breastfed their infants without supplementation.

  16. Radiosensitivity of the rough-skinned newt (Taricha granulosa)

    International Nuclear Information System (INIS)

    Willis, D.L.; Lappenbusch, W.L.

    1976-01-01

    Newts were collected locally and maintained unfed at 10 0 C both before and after irradiation. Whole-body exposures ranging up to 80 kR were given with 250 kVp x-rays, 300 kVp x-rays and 60 Co gamma-rays. The mean survival time-exposure curve was sigmoid, following the typical appearance of such curves for mammalian species. A dose-independent region extending from roughly 700 R to 5000 R was evident. A variety of skin lesions and depigmentation effects were noted at exposures above 250 R, whose time of appearance decreased with increasing dose. A total of 832 newts were used in this phase of the study. Assuming that hematopoietic damage was primarily responsible for deaths below 700 to 800 R, additional studies of hematopoiesis were initiated. The effect of a 650 R-exposure on liver weight, spleen weight and concentration of circulating red blood cells for 1 1 / 2 months postirradiation was assessed. The results indicate a progressive state of anemia. This was further substantiated by a study of 59 Fe incorporation by red blood cells at intervals up to 1 month for five exposures ranging up to 1000 R

  17. Effects of dietary antioxidant supplementation in cow’s feed, milk processing and storage on lutein content and sensory quality

    Science.gov (United States)

    In this work, we studied the lutein content in milk as affected by lutein supplementation in the absence and presence of common antioxidants, vitamin E (Vit E), tea polyphenols (TP) and ethoxyquin (EQ) in cow’s feed, and by subsequent pasteurization (HTST and UHT) and storage. Results showed that l...

  18. Role of lutein in alleviating the effects of electron beam radiation induced hematological and biochemical changes in Swiss albino mice

    International Nuclear Information System (INIS)

    Vidya, V.; Krishna, A.P.; Patil, Shrikant; Fernandes, Ronald

    2016-01-01

    Lutein is a naturally occurring xanthophyll pigment derived from α-carotene. It is abundantly present in Tagetes erecta L. (marigold) and also present in a few vegetables, fruits and in animal sources. Lutein was evaluated for its protective role in electron beam radiation induced damages in Swiss albino mice. The drug was optimized for its radioprotective activity

  19. Macular pigment density in relation to serum and adipose tissue concentrations of lutein and serum concentrations of zeaxanthin

    NARCIS (Netherlands)

    Broekmans, W.M.R.; Berendschot, T.T.J.R.; Klöpping-Ketelaars, I.A.A.; Vries, A.J. de; Goldbohm, R.A.; Tijburg, L.B.M.; Kardinaal, A.F.M.; Poppel, G. van

    2002-01-01

    Background: Macular pigment (MP), concentrated in the central area of the retina, contains the carotenoids lutein and zeaxanthin. A low MP density could be a risk factor for age-related macular degeneration. Little information is available regarding MP density in relation to serum lutein and

  20. Biosynthesis and the conjugation of magnetite nanoparticles with luteinizing hormone releasing hormone (LHRH)

    Energy Technology Data Exchange (ETDEWEB)

    Obayemi, J.D. [Department of Materials Science and Engineering, African University of Science and Technology (AUST) Abuja, Federal Capital Territory (Nigeria); Department of Materials Science and Engineering, Kwara State University, Malete, Kwara State (Nigeria); Dozie-Nwachukwu, S. [Department of Materials Science and Engineering, African University of Science and Technology (AUST) Abuja, Federal Capital Territory (Nigeria); Sheda Science and Technology Complex (SHESTCO) Abuja, Federal Capital Territory (Nigeria); Danyuo, Y. [Department of Materials Science and Engineering, African University of Science and Technology (AUST) Abuja, Federal Capital Territory (Nigeria); Department of Electronics and Electricals Engineering, Nigerian Turkish Nile University, Abuja (Nigeria); Odusanya, O.S. [Department of Materials Science and Engineering, African University of Science and Technology (AUST) Abuja, Federal Capital Territory (Nigeria); Sheda Science and Technology Complex (SHESTCO) Abuja, Federal Capital Territory (Nigeria); Anuku, N. [Department of Chemistry, Bronx Community College, New York, NY 10453 (United States); Princeton Institute of Science and Technology of Materials (PRISM), Princeton, NJ 08544 (United States); Malatesta, K. [Princeton Institute of Science and Technology of Materials (PRISM), Princeton, NJ 08544 (United States); Department of Mechanical and Aerospace Engineering, Princeton University, NJ 08544 (United States); Soboyejo, W.O., E-mail: soboyejo@princeton.edu [Department of Materials Science and Engineering, African University of Science and Technology (AUST) Abuja, Federal Capital Territory (Nigeria); Princeton Institute of Science and Technology of Materials (PRISM), Princeton, NJ 08544 (United States); Department of Mechanical and Aerospace Engineering, Princeton University, NJ 08544 (United States)

    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. - Highlights: • Biosynthesis of MNPs with clinically relevant sizes between 10 and 60 nm. • New insights into the effects of pH and processing time on nanoparticle shapes and sizes. • Successful conjugation of biosynthesized magnetite nanoparticles to LHRH ligands. • Conjugated BMNPs that are monodispersed with potential biomedical relevance. • Magnetic properties of biosynthesized MNPs suggest potential for MRI enhancement.

  1. Biosynthesis and the conjugation of magnetite nanoparticles with luteinizing hormone releasing hormone (LHRH)

    International Nuclear Information System (INIS)

    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. - Highlights: • Biosynthesis of MNPs with clinically relevant sizes between 10 and 60 nm. • New insights into the effects of pH and processing time on nanoparticle shapes and sizes. • Successful conjugation of biosynthesized magnetite nanoparticles to LHRH ligands. • Conjugated BMNPs that are monodispersed with potential biomedical relevance. • Magnetic properties of biosynthesized MNPs suggest potential for MRI enhancement

  2. 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

  3. Clinical trial of lutein in patients with retinitis pigmentosa receiving vitamin A treatment