Ganglioside inhibition of attachment and differentiation of cultured rat granulosa cells: interactions with fibronectin

The involvement of fibronectin in the attachment and differentiation of rat granulosa cells, cultured in a chemically defined medium, was investigated using the inhibitory properties of mixed brain gangliosides (MBGs) and highly purified disialoganglioside, GD1a. MBGs inhibited cell attachment to plastic culture surfaces in a concentration-dependent manner, with 0.1 mmol l-1 causing significantly decreased attachment between 0.5 and 24 h of incubation. Inhibition of attachment to a fibronectin-coated substratum was even greater. The inhibitory effect of MBGs was not caused by binding to the cell surface, but instead the inhibitory factor(s) were adsorbed on a surface of immobilized human plasma fibronectin, thereby preventing cell attachment to this surface. The inhibitory action of MBGs was also neutralized by the addition of soluble fibronectin. Furthermore, at least one component of MBGs, detected chemically following thin-layer chromatography, was directly shown to bind to human fibronectin. MBGs inhibited to varying degrees the follicle-stimulating hormone(FSH)-dependent responses: augmentation of cellular protein content, production of adenosine 3′,5′-cyclic monophosphate (cyclic AMP) and progestins (progesterone + 20 alpha-hydroxypregn-4-en-3-one + pregnenolone), and induction of aromatase activity. These inhibitory activities of MBGs could not be eliminated by adsorption on immobilized fibronectin or reversed by addition of soluble fibronectin, thus distinguishing these actions from the early inhibition of cell attachment. FSH-dependent responses were also inhibited by GD1a, while responses to stimulation by dibutyryl cyclic AMP plus 3-isobutyl-1-methyl xanthine were less affected by this ganglioside. These results suggest that gangliosides inhibit attachment of granulosa cells in culture by binding to fibronectin, whereas the inhibition of FSH-dependent differentiation occurs by other modes of action that are unrelated to the effects on cell adhesion.

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Inhibitory effect of Sphagnum palustre extract and its bioactive compounds on aromatase activity

Bangladesh Journal of Pharmacology ◽

10.3329/bjp.v11i3.26776 ◽

2016 ◽

Vol 11 (3) ◽

pp. 661

Author(s):

Hee Jeong Eom ◽

Yong Joo Park ◽

Hee Rae Kang ◽

Ha Ryong Kim ◽

In Jae Bang ◽

...

Keyword(s):

Video Clip ◽

Inhibitory Effect ◽

The Other ◽

Aromatase Activity ◽

Dependent Manner ◽

Inhibitory Effects ◽

Aromatase Inhibition ◽

Concentration Dependent Manner ◽

Cardiac Pain ◽

Sphagnum Palustre

Sphagnum palustre (a moss) has been traditionally used in Korea for the cure of several diseases such as cardiac pain and stroke. In this research, the inhibitory effect of S. palustre on aromatase (cytochrome P450 19, CYP19) activity was studied. [1β-3H] androstenedione was used as a substrate and incubated with S. palustre extract and recombinant human CYP19 in the presence of NADPH. S. palustre extract inhibited aromatase in a concentration-dependent manner (IC50 value: 36.4 ± 8.1 µg/mL). To elucidate the major compounds responsible for the aromatase inhibitory effects of S. palustre extract, nine compounds were isolated from the extract and tested for their inhibition of aromatase activity. Compounds 1, 6, and 7 displayed aromatase inhibition, while the inhibition by the other compounds was negligible.Video Clip<a href="https://youtube.com/v/n6xeo3RXJVY">Aromatase enzyme activity:</a> 4 min 16 sec

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Inhibition of aromatase activity in rat Sertoli cells by thyroid hormone

Journal of Endocrinology ◽

10.1677/joe.0.1400431 ◽

1994 ◽

Vol 140 (3) ◽

pp. 431-436 ◽

Cited By ~ 38

Author(s):

S Ulisse ◽

E A Jannini ◽

E Carosa ◽

D Piersanti ◽

F M Graziano ◽

...

Keyword(s):

Thyroid Hormone ◽

Cyclic Amp ◽

Sertoli Cells ◽

Inhibitory Effect ◽

Aromatase Activity ◽

Dependent Manner ◽

Maximal Dose ◽

Cyclic Amp Accumulation ◽

Order Of Magnitude ◽

Dose Dependent

Abstract Basal and FSH-induced aromatase activity in prepubertal rat Sertoli cells was inhibited by l-tri-iodothyronine (T3) in a time- and dose-dependent manner. The effect was evident only after 6 h of preincubation with T3 (10−7 m) and the half-maximal dose was 0·5 ±0·2 nm, which correlated with the Kd of the nuclear T3 receptor of rat Sertoli cells (Kd=1–2 nm). The effect was specific as judged by the lack of effect of the T3 analogue 3-iodo-l-thyrosine. The inhibitory effect of T3 was present over the entire range of FSH concentrations used (0·001–100 ng/ml). In T3-treated Sertoli cells, aromatase activity induced by 8-bromo-cyclic AMP was inhibited by the same order of magnitude as that of FSH, thus suggesting that the inhibitory effect of T3 was downstream from cyclic AMP formation. Furthermore, pretreatment of Sertoli cells cultures with T3 (24 h, 10−7 m) did not affect basal or FSH-induced extracellular cyclic AMP accumulation. This effect of T3 on rat Sertoli cell aromatase activity may be regarded as a part of the integrated mechanism by which thyroid hormone modulates the functions of the seminiferous epithelium. Journal of Endocrinology (1994) 140, 431–436

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Direct effects of bromocriptine on the steroidogenic capability of porcine granulosa cells

Acta Endocrinologica ◽

10.1530/acta.0.1260338 ◽

1992 ◽

Vol 126 (4) ◽

pp. 338-344 ◽

Cited By ~ 1

Author(s):

Shusaku Kamada ◽

Toshiro Kubota ◽

Makoto Taguchi ◽

Takeshi Aso

Keyword(s):

Direct Effect ◽

Granulosa Cells ◽

Inhibitory Effect ◽

Cell Number ◽

Dependent Manner ◽

Direct Effects ◽

Concentration Dependent Manner ◽

Ovarian Granulosa Cells ◽

Estradiol Secretion ◽

Porcine Granulosa Cells

The direct effects of bromocriptine on steroidogenesis were examined in cultured porcine granulosa cells. The following observations were made with bromocriptine: (1) It significantly increased the basal or FSH-stimulated secretion of progesterone in cultured porcine granulosa cells at concentrations exceeding 10−7 mol/l; (2) its inhibitory effect on basal estradiol secretion was demonstrated; (3) it did not influence cell number in cultured porcine granulosa cells; (4) it increased the extracellular accumulation of cAMP in a concentration-dependent manner; and (5) it did not induce a change in cytosolic free Ca2+ concentration. These findings suggest that bromocriptine exerts a direct effect on steroidogenesis in ovarian granulosa cells.

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Adhesion and differentiation of cultured rat granulosa cells: role of fibronectin

AJP Cell Physiology ◽

10.1152/ajpcell.1987.253.5.c625 ◽

1987 ◽

Vol 253 (5) ◽

pp. C625-C632 ◽

Cited By ~ 22

Author(s):

P. Morley ◽

D. T. Armstrong ◽

R. E. Gore-Langton

Keyword(s):

Granulosa Cell ◽

Granulosa Cells ◽

Inhibitory Activity ◽

Cell Attachment ◽

Follicle Stimulating Hormone ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

Cyclic Monophosphate ◽

Secretion Product

Because fibronectin is a major secretion product of rat granulosa cells in culture, we have investigated 1) the involvement of endogenous fibronectin in granulosa cell attachment, and 2) the consequences of inhibition of this attachment on follicle-stimulating hormone (FSH)-dependent differentiated responses. Attachment was significantly inhibited for up to 8 h in a concentration-dependent manner by antiserum to rat fibronectin, but not by nonimmune serum. Adsorption of antiserum on fibronectin or addition of exogenous fibronectin eliminated this inhibitory activity. Treatment with antiserum did not significantly alter the FSH-dependent production of adenosine 3',5'-cyclic monophosphate over 2 h or progestins over 48 h, while conversion of testosterone to 17 beta-estradiol over 48 h was suppressed by 60% in the presence of antiserum, regardless of antiserum adsorption on fibronectin. Results indicate that endogenous fibronectin is involved in substratum attachment of rat granulosa cells, but that attachment is not a requisite for FSH responsiveness.

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Modulation of differentiation of rat granulosa cells in vitro by interferon-γ

Journal of Endocrinology ◽

10.1677/joe.0.1330131 ◽

1992 ◽

Vol 133 (1) ◽

pp. 131-139 ◽

Cited By ~ 12

Author(s):

S. Xiao ◽

J. K. Findlay

Keyword(s):

Granulosa Cell ◽

Granulosa Cells ◽

Cell Function ◽

Inhibitory Effect ◽

Interferon Γ ◽

Detectable Effect ◽

Aromatase Activity ◽

Dependent Manner ◽

Ifn Γ

ABSTRACT The effects of recombinant rat interferon-γ (rRaIFN-γ) and rat IFN (RaIFN, a mixture of IFN-γ and -α) on basal and FSH-induced ovarian granulosa cell function were studied. Granulosa cells were harvested from diethylstilboestrol-treated immature rats and cultured (2 × 105 viable cells/well per 0·5 ml) in serumfree medium with or without treatment for 48 h. In the presence of FSH (20 ng/ml), rRaIFN-γ (10–1000 U/ml) significantly inhibited FSH-stimulated aromatase activity (76·4 ± 2·3% maximum inhibition compared with FSH treatment alone), inhibin (40·4 ± 3·7%), progesterone (47·7 ± 8·6%) and 20α-hydroxypregn-4-en-3-one (20α-OHP) (51·8±1·7%) production in a dose-dependent manner. Furthermore, rRaIFN-γ inhibited FSH- and forskolin (FSK; 30 μmol/l)-induced extracellular cAMP accumulation (46·0 ± 6·6% and 29·1 ± 7·3% respectively). The inhibitory effect of rRaIFN-γ on FSK-induced cAMP was accompanied by decreased FSK-induced aromatase activity, inhibin, progesterone and 20α-OHP production. rRaIFN-γ had no detectable effect on aromatase activity, progesterone production and 20α-OHP production in the absence of FSH, but significantly stimulated basal inhibin production by 1·5-fold. rRaIFN-γ alone also caused a small but significant increase in basal levels of cAMP. The timecourse studies showed that FSH-induced aromatase activity and inhibin production were consistently suppressed by rRaIFN-γ, FSH-induced progesterone and 20α-OHP were inhibited at 1 and 2 days and then stimulated on days 3, 4 and 5 relative to FSH alone. There was no difference in DNA content between treatment and non-treatment wells during 5 days of culture. RaIFN had similar effects to rRaIFN-γ. We conclude that IFN-γ can inhibit FSH-induced granulosa cell differentiation and that, in the absence of FSH, IFN-γ stimulated undifferentiated granulosa cells to produce more inhibin. The mechanism of its action is likely to involve changes in cAMP production. Journal of Endocrinology (1992) 133, 131–139

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Brassinolide Enhances the Level of Brassinosteroids, Protein, Pigments, and Monosaccharides in Wolffia arrhiza Treated with Brassinazole

Plants ◽

10.3390/plants10071311 ◽

2021 ◽

Vol 10 (7) ◽

pp. 1311

Author(s):

Magdalena Chmur ◽

Andrzej Bajguz

Keyword(s):

Plant Growth ◽

Growth And Development ◽

Inhibitory Effect ◽

Dependent Manner ◽

Plant Growth And Development ◽

Concentration Dependent Manner ◽

Spectrophotometric Methods ◽

Synthetic Inhibitor ◽

Wolffia Arrhiza ◽

First Time

Brassinolide (BL) represents brassinosteroids (BRs)—a group of phytohormones that are essential for plant growth and development. Brassinazole (Brz) is as a synthetic inhibitor of BRs’ biosynthesis. In the present study, the responses of Wolffia arrhiza to the treatment with BL, Brz, and the combination of BL with Brz were analyzed. The analysis of BRs and Brz was performed using LC-MS/MS. The photosynthetic pigments (chlorophylls, carotenes, and xanthophylls) levels were determined using HPLC, but protein and monosaccharides level using spectrophotometric methods. The obtained results indicated that BL and Brz influence W. arrhiza cultures in a concentration-dependent manner. The most stimulatory effects on the growth, level of BRs (BL, 24-epibrassinolide, 28-homobrassinolide, 28-norbrassinolide, catasterone, castasterone, 24-epicastasterone, typhasterol, and 6-deoxytyphasterol), and the content of pigments, protein, and monosaccharides, were observed in plants treated with 0.1 µM BL. Whereas the application of 1 µM and 10 µM Brz caused a significant decrease in duckweed weight and level of targeted compounds. Application of BL caused the mitigation of the Brz inhibitory effect and enhanced the BR level in duckweed treated with Brz. The level of BRs was reported for the first time in duckweed treated with BL and/or Brz.

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Adenosine A1 receptor-mediated inhibition of vasopressin action in inner medullary collecting duct

AJP Renal Physiology ◽

10.1152/ajprenal.1994.266.5.f791 ◽

1994 ◽

Vol 266 (5) ◽

pp. F791-F796 ◽

Cited By ~ 11

Author(s):

R. M. Edwards ◽

W. S. Spielman

Keyword(s):

Receptor Agonist ◽

Collecting Duct ◽

Basolateral Membrane ◽

Inhibitory Effect ◽

Dependent Manner ◽

Camp Accumulation ◽

Concentration Dependent Manner ◽

Cgs 21680 ◽

Camp Generation ◽

A1 Receptor

We examined the effects of adenosine and adenosine analogues on arginine vasopressin (AVP)-induced increases in osmotic water permeability (Pf; micron/s) and adenosine 3',5'-cyclic monophosphate (cAMP) accumulation in rat inner medullary collecting ducts (IMCDs). When added to the bath, the A1 receptor agonist N6-cyclohexyladenosine (CHA) produced a rapid and reversible inhibition of AVP-stimulated (10 pM) Pf (1,781 +/- 195 to 314 +/- 85 microns/s at 0.3 microM CHA; n = 9). The inhibitory effect of CHA was concentration dependent, with a 50% inhibitory concentration of 10 nM. The effect of CHA was inhibited by prior exposure of IMCDs to the A1 receptor antagonist 1,3-dipropylxanthine-8-cyclopentylxanthine (DP-CPX; 1 microM) or by preincubation with pertussis toxin. CHA had no effect on cAMP-induced increases in Pf. In addition to CHA, adenosine and the nonselective agonist 5'-(N-ethylcarboxamido)-adenosine (NECA) inhibited AVP-dependent Pf by > or = 70%, whereas the A2 receptor agonist CGS-21680 had no effect. Luminal adenosine (0.1 mM) had no effect on basal or AVP-stimulated Pf. CHA, NECA, and adenosine but not CGS-21680 inhibited AVP-stimulated cAMP accumulation in a concentration-dependent manner (50% inhibitory concentrations 0.1–300 nM). The inhibitory effect of CHA on AVP-stimulated cAMP accumulation was attenuated by DPCPX. We conclude that adenosine, acting at the basolateral membrane, inhibits AVP action in the IMCD via interaction with A1 receptors. The inhibition occurs proximal to cAMP generation and likely involves an inhibitory G protein.

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Fibrinogen binding to the integrin αIIbβ3 modulates store-mediated calcium entry in human platelets

Blood ◽

10.1182/blood.v97.9.2648 ◽

2001 ◽

Vol 97 (9) ◽

pp. 2648-2656 ◽

Cited By ~ 22

Author(s):

Juan A. Rosado ◽

Else M. Y. Meijer ◽

Karly Hamulyak ◽

Irena Novakova ◽

Johan W. M. Heemskerk ◽

...

Keyword(s):

Actin Cytoskeleton ◽

Actin Polymerization ◽

Adenosine Diphosphate ◽

Inhibitory Effect ◽

Human Platelets ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

Integrin Αiibβ3 ◽

Fibrinogen Binding

Abstract Effects of the occupation of integrin αIIbβ3 by fibrinogen on Ca++signaling in fura-2–loaded human platelets were investigated. Adding fibrinogen to washed platelet suspensions inhibited increases in cytosolic [Ca++] concentrations ([Ca++]i) evoked by adenosine diphosphate (ADP) and thrombin in a concentration-dependent manner in the presence of external Ca++ but not in the absence of external Ca++ or in the presence of the nonselective cation channel blocker SKF96365, indicating selective inhibition of Ca++entry. Fibrinogen also inhibited store-mediated Ca++ entry (SMCE) activated after Ca++ store depletion using thapsigargin. The inhibitory effect of fibrinogen was reversed if fibrinogen binding to αIIbβ3 was blocked using RDGS or abciximab and was absent in platelets from patients homozygous for Glanzmann thrombasthenia. Fibrinogen was without effect on SMCE once activated. Activation of SMCE in platelets occurs through conformational coupling between the intracellular stores and the plasma membrane and requires remodeling of the actin cytoskeleton. Fibrinogen inhibited actin polymerization evoked by ADP or thapsigargin in control cells and in cells loaded with the Ca++ chelator dimethyl BAPTA. It also inhibited the translocation of the tyrosine kinase p60src to the cytoskeleton. These results indicate that the binding of fibrinogen to integrin αIIbβ3 inhibits the activation of SMCE in platelets by a mechanism that may involve modulation of the reorganization of the actin cytoskeleton and the cytoskeletal association of p60src. This action may be important in intrinsic negative feedback to prevent the further activation of platelets subjected to low-level stimuli in vivo.

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Corticotropin-Releasing Factor Inhibits Luteinizing Hormone-Stimulated P450c17 Gene Expression and Androgen Production by Isolated Thecal Cells of Human Ovarian Follicles1

The Journal of Clinical Endocrinology & Metabolism ◽

10.1210/jcem.83.2.4546 ◽

1998 ◽

Vol 83 (2) ◽

pp. 448-452

Author(s):

H. F. Erden ◽

I. H. Zwain ◽

H. Asakura ◽

S. S. C. Yen

Keyword(s):

Gene Expression ◽

Granulosa Cells ◽

Inhibitory Effect ◽

Corticotropin Releasing Factor ◽

Mrna Levels ◽

Dependent Manner ◽

Messenger Ribonucleic Acid ◽

Estrogen Production ◽

Thecal Cells ◽

Crf System

Recently, we reported that the thecal compartment of the human ovary contains a CRF system replete with gene expression and protein for corticotropin-releasing factor (CRF), CRF-Receptor 1 (CRF-R1), and the blood-derived high affinity CRF-binding protein (CRF-BP). Granulosa cells are devoid of the CRF system. The parallel increases in intensity of CRF, CRF-R1, and 17α-hydroxylase messenger ribonucleic acid (mRNA) and proteins in thecal cells with follicular maturation suggest that the intraovarian CRF system may play an autocrine role regulating androgen biosynthesis, with a downstream effect on estrogen production by granulosa cells. The functionality of the ovarian CRF system may be conditioned by the relative presence of plasma-derived CRF-BP by virtue of its localization of protein, but not transcript in thecal cells and its ability to compete with CRF for the CRF receptor. To further these findings, in the present study we have examined the effect of CRF on LH-stimulated 17α-hydroxylase (P450c17) gene expression and androgen production by isolated thecal cells from human ovarian follicles (11–13 mm). During the 48-h culture, addition of LH (10 ng/mL) to the medium increased by 5- and 6-fold dehydroepiandrosterone and androstenedione production by thecal cells. Remarkably, the LH-stimulated, but not basal, androgen production was inhibited by CRF in a time- and dose-dependent manner. The half-maximal (ID50) effect dose of CRF occurred at 5 × 10−8 mol/L, and at a maximal concentration of 10−6 mol/L, CRF completely inhibited LH-stimulated androgen production. This inhibitory effect of CRF became evident at 12 h (45%), and by 24 h the effect was more pronounced, with a 70% reduction from baseline. As determined by Northern analyses, CRF dose dependently decreased LH-stimulated P450c17 mRNA levels, with a maximal inhibition of 85% P450c17 gene expression at a CRF concentration of 10−6 mol/L. With the addition of 10−6 mol/L of the antagonist α-helical CRF-(9–41), the inhibitory effect of CRF was partially reversed for both P450c17 mRNA (75%) and androgen production (50%), indicating the CRF-R1-mediated event. In conclusion, the present study demonstrated a potent inhibitory effect of CRF on LH-stimulated dehydroepiandrosterone and androstenedione production that appears to be mediated through the reduction of P450c17 gene expression. Thus, the ovarian CRF system may function as autocrine regulators for androgen biosynthesis in the thecal cell compartment to maintain optimal substrate for estrogen biosynthesis by granulosa cells. Further studies to define the role of CRF-BP in the endocrine modulation of the intraovarian CRF system are needed.

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Thrombin activatable fibrinolysis inhibitor (TAFI) affects fibrinolysis in a plasminogen activator concentration-dependent manner

Thrombosis and Haemostasis ◽

10.1160/th03-06-0377 ◽

2004 ◽

Vol 91 (03) ◽

pp. 473-479 ◽

Cited By ~ 15

Author(s):

Ana Guimarães ◽

Dingeman Rijken

Keyword(s):

Plasminogen Activator ◽

In Vitro Study ◽

Inhibitory Effect ◽

Retardation Factor ◽

Clot Lysis ◽

Dependent Manner ◽

Plasma Clot ◽

Concentration Dependent Manner ◽

Plasmin Inhibitor

SummaryTAFIa was shown to attenuate fibrinolysis. In our in vitro study, we investigated how the inhibitory effect of TAFIa depended on the type and concentration of the plasminogen activator (PA). We measured PA-mediated lysis times of plasma clots under conditions of maximal TAFI activation by thrombin-thrombomodulin in the absence and presence of potato carboxypeptidase inhibitor. Seven different PAs were compared comprising both tPA-related (tPA, TNK-tPA, DSPA), bacterial PA-related (staphylokinase and APSAC) and urokinase-related (tcu-PA and k2tu-PA) PAs. The lysis times and the retardation factor were plotted against the PA concentration. The retardation factor plots were bell-shaped. At low PA concentrations, the retardation factor was low, probably due to the limited stability of TAFIa. At intermediate PA concentrations the retardation factor was maximal (3-6 depending on the PA), with TNK-tPA, APSAC and DSPA exhibiting the strongest effect. At high PA concentrations, the retardation factor was again low, possibly due to inactivation of TAFIa by plasmin or to a complete conversion of glu-plasminogen into lys-plasminogen. Using individual plasmas with a reduced plasmin inhibitor activity (plasmin inhibitor Enschede) the bell-shaped curve of the retardation factor shifted towards lower tPA and DSPA concentrations, but the height did not decrease. In conclusion, TAFIa delays the lysis of plasma clots mediated by all the plasminogen activators tested. This delay is dependent on the type and concentration of the plasminogen activator, but not on the fibrin specificity of the plasminogen activator. Furthermore, plasmin inhibitor does not play a significant role in the inhibition of plasma clot lysis by TAFI.

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