scholarly journals Effects of the mycotoxin deoxynivalenol on steroidogenesis and apoptosis in granulosa cells

Reproduction ◽  
2015 ◽  
Vol 149 (6) ◽  
pp. 555-561 ◽  
Author(s):  
Hilda M Guerrero-Netro ◽  
Younès Chorfi ◽  
Christopher A Price
Keyword(s):  
Granulosa Cell ◽  
Immune Cells ◽  
Culture System ◽  
Cell Function ◽  
Reproductive Function ◽  
Mrna Levels ◽  
Progesterone Secretion ◽  
Ribotoxic Stress Response ◽  
Jnk Activation ◽  
Dose Dependent

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.

scholarly journals Adenosine 5′-Monophosphate-Activated Protein Kinase Regulates Progesterone Secretion in Rat Granulosa Cells

Endocrinology ◽  
2005 ◽  
Vol 146 (10) ◽  
pp. 4500-4513 ◽  
Author(s):  
Lucie Tosca ◽  
Pascal Froment ◽  
Patricia Solnais ◽  
Pascal Ferré ◽  
Fabienne Foufelle ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Granulosa Cells ◽  
Dominant Negative ◽  
Detectable Effect ◽  
Mrna Levels ◽  
Progesterone Secretion ◽  
Igf I ◽  
Amp Activated Protein Kinase ◽  
Acetyl Coenzyme A Carboxylase ◽  
Dose Dependent

The AMP-activated protein kinase (AMPK) is a major regulator of energy metabolism involved in fatty acid and cholesterol synthesis. In the ovary, cholesterol plays a key role in steroid production. We report the presence of AMPK in rat ovaries, and we have investigated its role in granulosa cells. We show using RT-PCR and Western blot that the mRNAs for the α1/2 and β1/2 subunits and the proteins are found in the ovaries. Immunohistochemistry localized the α1 AMPK subunit in granulosa cells, corpus luteum, and oocyte and less abundantly in theca cells. Treatment with 1 mm 5-amino-imidazole-4-carboxyamide-1-β-d-ribofuranoside (AICAR), an activator of AMPK, increased dose-dependent and time-dependent phosphorylation of AMPKα1 on Thr172 in primary granulosa cells. Simultaneously, phosphorylation of acetyl-coenzyme A carboxylase at Ser79 was also increased. AICAR treatment for 48 h halved progesterone secretion, 3β-HSD protein and mRNA levels, and phosphorylation of both basal MAPK ERK1/2 and p38 and in response to IGF-I and/or FSH in granulosa cells. AICAR treatment (1 mm) had no detectable effect on basal and FSH- and/or IGF-I-induced estradiol production and on granulosa cell proliferation or viability. Adenovirus-mediated expression of dominant negative AMPK totally abolished the effects of AICAR on progesterone secretion, 3β-HSD protein production, and MAPK ERK1/2 and p38 phosphorylation. Moreover, we showed using specific in- hibitors of ERK1/2 and p38 MAPK that the MAPK ERK1/2 and not p38 is involved in progesterone secretion and 3β-HSD expression, strongly suggesting that the activation of AMPK in response to AICAR reduces progesterone production through the MAPK ERK1/2 signaling pathway in rat granulosa cells.

scholarly journals MicroRNA-224 Is Involved in Transforming Growth Factor-β-Mediated Mouse Granulosa Cell Proliferation and Granulosa Cell Function by Targeting Smad4

2010 ◽  
Vol 24 (3) ◽  
pp. 540-551 ◽  
Author(s):  
Guidong Yao ◽  
Mianmian Yin ◽  
Jie Lian ◽  
Hui Tian ◽  
Lin Liu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Granulosa Cell ◽  
Transforming Growth Factor ◽  
Cell Function ◽  
Ectopic Expression ◽  
Follicular Development ◽  
Transforming Growth Factor Β ◽  
Type I ◽  
Mrna Levels ◽  
Tgf Β1

Abstract Many members of the TGF-β superfamily are indicated to play important roles in ovarian follicular development, such as affecting granulosa cell function and oocyte maturation. Abnormalities associated with TGF-β1 signaling transduction could result in female infertility. MicroRNAs (miRNAs), as small noncoding RNAs, were recently found to regulate gene expression at posttranscriptional levels. However, little is known about the role of miRNAs in TGF-β-mediated granulosa cell proliferation and granulosa cell function. In this study, the miRNA expression profiling was identified from TGF-β1-treated mouse preantral granulosa cells (GCs), and three miRNAs were found to be significantly up-regulated and 13 miRNAs were down-regulated. Among up-regulated miRNAs, miR-224 was the second most significantly elevated miRNA. This up-regulation was attenuated by treatment of GCs with SB431542 (an inhibitor of TGFβ superfamily type I receptors, thus blocking phosphorylation of the downstream effectors Smad2/3), indicating that miR-224 expression was regulated by TGF-β1/Smads pathway. The ectopic expression of miR-224 can enhance TGF-β1-induced GC proliferation through targeting Smad4. Inhibition of endogenous miR-224 partially suppressed GC proliferation induced by TGF-β1. In addition, both miR-224 and TGF-β1 can promote estradiol release from GC, at least in part, through increasing CYP19A1 mRNA levels. This is the first demonstration that miRNAs can control reproductive functions resulting in promoting TGF-β1-induced GC proliferation and ovarian estrogen release. Such miRNA-mediated effects could be potentially used for regulation of reproductive processes or for treatment of reproductive disorders.

Effects of inhibin-related peptides and oestradiol on androstenedione and progesterone secretion by bovine theca cells in vitro

1995 ◽  
Vol 145 (3) ◽  
pp. 491-500 ◽  
Author(s):  
J H M Wrathall ◽  
P G Knight
Keyword(s):  
Granulosa Cell ◽  
Thecal Cell ◽  
Theca Cells ◽  
Progesterone Secretion ◽  
Pretreatment Period ◽  
Direct Contrast ◽  
Primary Monolayer ◽  
Dose Dependent

Abstract Primary monolayer cultures of bovine theca cells isolated from pooled ovarian follicles (3–10 mm diameter) were used to examine the effects of various granulosa cell-derived substances on basal and luteinizing hormone (LH)-induced androgen and progesterone secretion. After an overnight pretreatment period, cells were incubated with a range of treatments including LH, oestradiol-17β, inhibin, activin and follistatin. Media were collected after 48 h and assessment of androstenedione and progesterone secretion made by radioimmunoassay. Addition of LH (5–50 ng/ml) to the cells resulted in a dose-dependent stimulation of both androstenedione (2·5-to 3-fold rise; P<0·01) and progesterone (∼ 1·6-fold rise; P<0·001) production. Secretion of androstenedione was also raised (up to 5-fold; P<0·001) by addition of oestradiol-17β (0·3–300 ng/ml), whilst levels of the androgen in the presence of both LH (20 ng/ml) and oestradiol (300 ng/ml) were up to 12-fold higher (P<0·001) than control values. In contrast, oestradiol treatment inhibited by up to 50% both basal (P<0·001) and LH-stimulated (P<0·001) secretion of progesterone. Exposure of cells to purified bovine inhibin (5–125 ng/ml) consistently raised androstenedione secretion by up to 42% over basal levels (P<0·001). Inhibin also enhanced both LH-stimulated (∼20%; P<0·001) and oestradiol-stimulated (∼20%; P<0·05) secretion of androstenedione. In direct contrast, treatment of theca cells with human recombinant activin-A (1–50 ng/ml) inhibited both LH-stimulated (∼50%; P<0·001) and oestradiol-stimulated (∼30%; P<0·005) androstenedione secretion. Activin also reversed the positive effect of inhibin on basal (P<0·01), LH-stimulated (P<0·001) and oestradiol-stimulated (P<0·001) androstenedione secretion, though activin alone did not affect basal steroid output. Simultaneous addition of human recombinant follistatin reversed the inhibitory effects of activin on LH- and oestradiol-induced androstenedione secretion but did not modify the effects of inhibin. Follistatin alone did not alter either basal or LH-stimulated androstenedione output. Neither basal nor LH-stimulated secretion of progesterone were consistently affected by inhibin, activin or follistatin. As well as confirming the stimulatory effects of both LH and oestradiol on bovine thecal cell androgen production, these observations are indicative of opposing intrafollicular paracrine roles for granulosa cell-derived inhibin and activin in modulating thecal cell responses to gonadotrophins and steroids in the bovine ovary. Though inhibin and oestradiol had qualitatively similar effects in promoting thecal androgen secretion, the magnitude of the response to oestradiol was much greater. The results also support an intrafollicular role of follistatin as a binding protein capable of neutralizing the effect of activin, but not inhibin, on thecal androgen production. Journal of Endocrinology (1995) 145, 491–500

INHIBITION OF FSH-STIMULATED GRANULOSA CELL FUNCTION BY A SYNTHETIC FRAGMENT OF THE PORCINE INHIBIN α-SUBUNIT: EVIDENCE FOR INVOLVEMENT OF GnRH RECEPTORS

1987 ◽  
Vol 113 (2) ◽  
pp. R3-R5 ◽  
Author(s):  
S.G. Hillier ◽  
C.G. Tsonis ◽  
E.J. Wickings ◽  
K.A. Eidne
Keyword(s):  
Granulosa Cell ◽  
Culture Medium ◽  
Cell Function ◽  
Specific Binding ◽  
Female Rat ◽  
Terminal Sequence ◽  
Α Subunit ◽  
Gnrh Receptors ◽  
Synthetic Peptide Fragment ◽  
Dose Dependent

ABSTRACT The bioactivity of a synthetic peptide fragment which mimics the N-terminal sequence of the 134-amino-acid porcine Inhibin α-subunit (pl- α1-26-Gly27Tyr28-OH) was tested and compared with the bioactivity of GnRH in rat granulosa cell cultures. Granulosa cells from immature female rat ovaries were cultured with hFSH and testosterone to stimulate the production of cyclic AMP, progesterone and oestradiol. Addition of pl- α1-26-Gly27Tyr28-OH to the culture medium caused a dose-dependent suppression of all three parameters (ID50 700-1,000 nmol/l). GnRH caused similar but higher-potency inhibition (ID50 2-4 nmol/l). Suppression of granulosa cell function by both peptides was fully reversible by a synthetic GnRH antagonist. Moreover, specific binding of the porcine inhibin fragment to ovarian GnRH receptors was demonstrated by radioreceptor assay. This is evidence that the porcine inhibin α-subunit fragment suppresses FSH-induced rat granulosa cell function via a mechanism of action similar to that of GnRH.

Bovine granulosa cell culture for assessment of potency and specificity of antibodies to pregnant mares' serum gonadotrophin

1996 ◽  
Vol 134 (4) ◽  
pp. 497-500 ◽  
Author(s):  
Mehmet Kuran ◽  
Peter J Broadbent ◽  
JS Morley Hutchinson
Keyword(s):  
Cell Culture ◽  
Granulosa Cell ◽  
Granulosa Cells ◽  
Culture System ◽  
Dependent Manner ◽  
Progesterone Production ◽  
Neutralizing Capacity ◽  
Dose Dependent

Kuran M, Broadbent PJ, Hutchinson JSM. Bovine granulosa cell culture for assessment of potency and specificity of antibodies to pregnant mares' serum gonadotrophin. Eur J Endocrinol 1996;134:497–500. ISSN 0804–4643 Antibodies to pregnant mares' serum gonadotrophin (PMSG) neutralize the effect of PMSG in vivo and increase the number of transferable embryos when administered at the optimum time relative to the preovulatory luteinizing hormone (LH) surge in PMSG-stimulated cows. The objective of the present study was to investigate the possible use of bovine granulosa cells in a serum-free culture system as a bioassay for antibodies to PMSG. Granulosa cells (2–3 × 105 viable cells) were cultured with varying doses of PMSG and/or an anti-PMSG for 4 days. Whilst progesterone production (ng/μg DNA) of granulosa cells was stimulated by PMSG (p < 0.01) in a dose-dependent manner, increasing amounts of anti-PMSG neutralized (p < 0.01) this stimulatory effect of either follicle-stimulating hormone or LH on progesterone production of bovine granulosa cells in vitro. The bovine granulosa cell culture system is a potential in vitro bioassay method for testing the specificity and the neutralizing capacity of different anti-PMSG preparations. Mehmet Kuran, Ondokuz Mayis Universitesi, Ziraat Fakultesi, Zootekni Bolumu, 55149 Samsun, Turkey

MiR-493 Induces Cytotoxic Autophagy in Prostate Cancer Cells through Regulation on PHLPP2

2020 ◽  
Vol 21 (14) ◽  
pp. 1451-1456 ◽  
Author(s):  
Jun Deng ◽  
Ming Ma ◽  
Wei Jiang ◽  
Liangliang Zheng ◽  
Suping Cui
Keyword(s):  
Prostate Cancer ◽  
Protein Expression ◽  
Cell Function ◽  
Mrna Levels ◽  
Prostate Cancer Cells ◽  
Potent Inducer ◽  
Qrt Pcr ◽  
Autophagy Gene ◽  
The Relationship ◽  
Cancer Inhibition

Background: MiR-493 promotes the proliferation of prostate cancer (PC) cells by targeting PHLPP2. We aimed to explore the relationship between miR-493 and autophagy in PC. Methods: qRT-PCR and western blotting were used to determine the mRNA levels and protein expression of miR-493, PHLPP2, autophagy gene BECN1 and ATG7 in PC cells. The autophagy gene expression was determined after PC cells transfected with miR-493 precursor or PHLPP2 precursor. Corresponding changes of autophagy phenotype and PC cell function were also studied. Results: The mRNA levels and protein expression of miR-493, PHLPP2, BECN1 and ATG7 in PC cells were significantly decreased in PC cells. Overexpression of miR-493 or PHLPP2 markedly upregulated the expression levels of BECN1 and ATG7 in PC cells. Overexpression of miR-493 and PHLPP2 markedly promoted autophagy, and inhibited the invasion and cloning formation of PC cells. Conclusion: MiR-493 is a potent inducer of cytotoxic autophagy that leads to prostate cancer inhibition by regulating on PHLPP2.

scholarly journals Sialic Acids and Their Influence on Human NK Cell Function

Cells ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 263
Author(s):  
Philip Rosenstock ◽  
Thomas Kaufmann
Keyword(s):  
Nk Cells ◽  
Immune Cells ◽  
Cell Function ◽  
Nk Cell ◽  
Sialic Acids ◽  
Innate Immune ◽  
Target Cells ◽  
Carbon Backbone ◽  
Nk Cell Receptors ◽  
Cell Inhibition

Sialic acids are sugars with a nine-carbon backbone, present on the surface of all cells in humans, including immune cells and their target cells, with various functions. Natural Killer (NK) cells are cells of the innate immune system, capable of killing virus-infected and tumor cells. Sialic acids can influence the interaction of NK cells with potential targets in several ways. Different NK cell receptors can bind sialic acids, leading to NK cell inhibition or activation. Moreover, NK cells have sialic acids on their surface, which can regulate receptor abundance and activity. This review is focused on how sialic acids on NK cells and their target cells are involved in NK cell function.

scholarly journals Glucolipotoxicity and GLP-1 secretion

2021 ◽  
Vol 9 (1) ◽  
pp. e001905
Author(s):  
Jung-Hee Hong ◽  
Dae-Hee Kim ◽  
Moon-Kyu Lee
Keyword(s):  
Glucose Transporter ◽  
Cell Function ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Gene Expressions ◽  
Simultaneous Treatment ◽  
L Cells ◽  
Triglyceride Accumulation

IntroductionThe concept of glucolipotoxicity refers to the combined, deleterious effects of elevated glucose and/or fatty acid levels.Research design and methodsTo investigate the effects of chronic glucolipotoxicity on glucagon-like peptide-1-(7-36) amide (GLP-1) secretion, we generated glucolipotoxic conditions in human NCI-H716 enteroendocrine cells using either 5 or 25 mM glucose with or without 500 µM palmitate for 72 hours. For in vivo study, we have established a chronic nutrient infusion model in the rat. Serial blood samples were collected for 2 hours after the consumption of a mixed meal to evaluate insulin sensitivity and β-cell function.ResultsChronic glucolipotoxic conditions decreased GLP-1 secretion and the expressions of pCREB, pGSK3β, β-catenin, and TCF7L2 in NCI-H716 cells. Glucolipotoxicity conditions reduced glucose transporter expression, glucose uptake, and nicotinamide-adenine dinucleotide phosphate (NADPH) levels in L-cells, and increased triglyceride accumulation. In contrast, PPARα and ATP levels were reduced, which correlated well with decreased levels of SUR1 and Kir6.2, cAMP contents and expressions of pCAMK2, EPAC and PKA. We also observed an increase in reactive oxygen species production, UCP2 expression and Complex I activity. Simultaneous treatment with insulin restored the GLP-1 secretion. Glucolipotoxic conditions decreased insulin secretion in a time-dependent manner in INS-1 cells, which was recovered with exendin-4 cotreatment. Glucose and SMOFlipid infusion for 6 hours decreased GLP-1 secretion and proglucagon mRNA levels as well as impaired the glucose tolerance, insulin and C-peptide secretion in rats.ConclusionThese results provide evidence for the first time that glucolipotoxicity could affect GLP-1 secretion through changes in glucose and lipid metabolism, gene expressions, and proglucagon biosynthesis and suggest the interrelationship between glucolipotoxicities of L-cells and β-cells which develops earlier than that of L-cells.

scholarly journals 601 Development of improved small molecule STING agonists suitable for systemic administration

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A636-A636
Author(s):  
Maciej Rogacki ◽  
Stefan Chmielewski ◽  
Magdalena Zawadzka ◽  
Jolanta Mazurek ◽  
Katarzyna Wnuk-Lipińska ◽  
...  
Keyword(s):  
Tumor Growth ◽  
Small Molecule ◽  
Immune Cells ◽  
Systemic Administration ◽  
Cytokine Release ◽  
Major Player ◽  
Human Immune ◽  
New Generation ◽  
Dose Dependent ◽  
Complete Tumor

BackgroundStimulator of Interferon Genes (STING) is a major player in the activation of robust innate immune response leading to initiation and enhancement of tumor-specific adaptive immunity. Several clinical and pre-clinical programs have shown that activation of the STING pathway triggers immune-mediated antitumor response. Although vast majority of programs focus on development of analogues of the endogenous STING ligands, their chemical nature and stability often limit their use to local administration. Herein, we present recent results from the development of our selective non-nucleotide, non-macrocyclic, small molecule direct STING agonists, suitable for systemic administration, characterized by improved activity in human immune cells.MethodsBinding to recombinant STING protein was examined using FTS, MST, FP and crystallography studies. Phenotypic screen was performed in THP-1 Dual reporter cells. Mouse bone marrow-derived dendritic cells (BMDC) were obtained from C57BL/6 mice and differentiated with mIL-4 and mGM-CSF. STING agonists were administered into BALB/c mice and cytokine release was measured in plasma. Additionally, mice were inoculated with CT26 murine colon carcinoma or EMT6 murine breast carcinoma cells and the compound was administered, followed by the regular tumor growth and body weight monitoring.ResultsRyvu’s small-molecule agonists demonstrate strong binding affinity to recombinant STING proteins across all tested species. The compounds bind to all human STING protein variants and trigger pro-inflammatory cytokine release from human immune cells regardless of the STING haplotype. Moreover, new generation of developed agonists show significantly improved binding to human protein as well as in vitro activity on human cells. Systemic, intravenous in vivo administration leads to a dose-dependent upregulation of STING-dependent pro-inflammatory cytokines, which results in a dose-dependent antitumor efficacy observed in CT26 and EMT6 mouse cancer models, leading to complete tumor remissions in all treated animals. Furthermore, observed efficacy is accompanied by development of a lasting immunological response demonstrated by lack of tumor engraftment or a delayed tumor growth in cured animals challenged with repeated inoculation of cancer cells.ConclusionsNew generation Ryvu’s STING agonists are strong and selective activators of STING-dependent signaling in both mouse and human immune cells promoting anti-tumor immunity. Treatment with Ryvu’s small-molecule STING agonists leads to engagement of the immune system which results in a complete tumor remission and development of immunological memory of the cancer antigens. The compounds show good selectivity and ADME properties enabling development for systemic administration. In addition developed compounds maintain small functional handles amenable to linker attachment making the series suitable for versatile development as single agents, for combinations with immunotherapies or as targeted agents.

scholarly journals Maternal Neutrophil Depletion Fails to Avert Systemic Lipopolysaccharide-Induced Early Pregnancy Defects in Mice

2021 ◽  
Vol 22 (15) ◽  
pp. 7932
Author(s):  
Sourav Panja ◽  
John T. Benjamin ◽  
Bibhash C. Paria
Keyword(s):  
Early Pregnancy ◽  
Normal Pregnancy ◽  
Mrna Levels ◽  
Interesting Approach ◽  
Blastocyst Implantation ◽  
Neutrophil Depletion ◽  
Underlying Mechanisms ◽  
Myd88 Signaling ◽  
Induced Defects ◽  
Dose Dependent

Maternal infection-induced early pregnancy complications arise from perturbation of the immune environment at the uterine early blastocyst implantation site (EBIS), yet the underlying mechanisms remain unclear. Here, we demonstrated in a mouse model that the progression of normal pregnancy from days 4 to 6 induced steady migration of leukocytes away from the uterine decidual stromal zone (DSZ) that surrounds the implanted blastocyst. Uterine macrophages were found to be CD206+ M2-polarized. While monocytes were nearly absent in the DSZ, DSZ cells were found to express monocyte marker protein Ly6C. Systemic endotoxic lipopolysaccharide (LPS) exposure on day 5 of pregnancy led to: (1) rapid (at 2 h) induction of neutrophil chemoattractants that promoted huge neutrophil infiltrations at the EBISs by 24 h; (2) rapid (at 2 h) elevation of mRNA levels of MyD88, but not Trif, modulated cytokines at the EBISs; and (3) dose-dependent EBIS defects by day 7 of pregnancy. Yet, elimination of maternal neutrophils using anti-Ly6G antibody prior to LPS exposure failed to avert LPS-induced EBIS defects allowing us to suggest that activation of Tlr4-MyD88 dependent inflammatory pathway is involved in LPS-induced defects at EBISs. Thus, blocking the activation of the Tlr4-MyD88 signaling pathway may be an interesting approach to prevent infection-induced pathology at EBISs.

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