Chemical Inhibitors of Dynamin Exert Differential Effects in VEGF Signaling

VEGFR2 is the main receptor and mediator of the vasculogenic and angiogenic activity of VEGF. Activated VEGFR2 internalizes through clathrin-mediated endocytosis and macropinocytosis. As dynamin is a key regulator of the clathrin pathway, chemical inhibitors of dynamin are commonly used to assess the role of the clathrin route in receptor signaling. However, drugs may also exert off-target effects. Here, we compare the effects of three dynamin inhibitors, dynasore, dyngo 4a and dynole, on VEGFR2 internalization and signaling. Although these drugs consistently inhibit clathrin-mediated endocytosis of both transferrin (a typical cargo of this route) and VEGFR2, surprisingly, they exert contradictory effects in receptor signaling. Thus, while dynasore has no effect on phosphorylation of VEGFR2, the other two drugs are strong inhibitors. Furthermore, although dyngo does not interfere with phosphorylation of Akt, dynasore and dynole have a strong inhibitory effect. These inconsistent effects suggest that the above dynamin blockers, besides inhibiting dynamin-dependent endocytosis of VEGFR2, exert additional inhibitory effects on signaling that are independent of endocytosis; i.e., they are due to off-target effects. Using a recently developed protocol, we comparatively validate the specificity of two endocytic inhibitors, dynasore and EIPA. Our findings highlight the importance of assessing whether the effect of an endocytic drug on signaling is specifically due to its interference with endocytosis or due to off-targets.

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Amphetamine-Decreased Progesterone and Estradiol Release in Rat Granulosa Cells: The Regulatory Role of cAMP- and Ca2+-Mediated Signaling Pathways

Biomedicines ◽

10.3390/biomedicines9050493 ◽

2021 ◽

Vol 9 (5) ◽

pp. 493

Author(s):

Chung-Yu Chen ◽

Chien-Rung Chen ◽

Chiao-Nan Chen ◽

Paulus S. Wang ◽

Toby Mündel ◽

...

Keyword(s):

Granulosa Cells ◽

Prostaglandin F2α ◽

Inhibitory Effect ◽

Inhibitory Effects ◽

Steroidogenic Enzyme ◽

Estradiol Secretion ◽

Basal Release ◽

Ca2 Channels

The purpose of this study is to evaluate the amphetamine effects on progesterone and estradiol production in rat granulosa cells and the underlying cellular regulatory mechanisms. Freshly dispersed rat granulosa cells were cultured with various test drugs in the presence of amphetamine, and the estradiol/progesterone production and the cytosolic cAMP level were measured. Additionally, the cytosolic-free Ca2+ concentrations ([Ca2+]i) were measured to examine the role of Ca2+ influx in the presence of amphetamine. Amphetamine in vitro inhibited both basal and porcine follicle-stimulating hormone-stimulated estradiol/progesterone release, and amphetamine significantly decreased steroidogenic enzyme activities. Adding 8-Bromo-cAMP did not recover the inhibitory effects of amphetamine on progesterone and estradiol release. H89 significantly decreased progesterone and estradiol basal release but failed to enhance a further amphetamine inhibitory effect. Amphetamine was capable of further suppressing the release of estradiol release under the presence of nifedipine. Pretreatment with the amphetamine for 2 h decreased the basal [Ca2+]i and prostaglandin F2α-stimulated increase of [Ca2+]i. Amphetamine inhibits progesterone and estradiol secretion in rat granulosa cells through a mechanism involving decreased PKA-downstream steroidogenic enzyme activity and L-type Ca2+ channels. Our current findings show that it is necessary to study the possibility of amphetamine perturbing reproduction in females.

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035. THE ROLE OF THE SERTOLI CELL IN REGULATING SPERMATOGENESIS, IMMUNE RESPONSES AND INFLAMMATORY DISEASE: MULTIPLE FUNCTIONS, COMMON MECHANISMS?

Reproduction Fertility and Development ◽

10.1071/srb09abs035 ◽

2009 ◽

Vol 21 (9) ◽

pp. 8

Author(s):

M. P. Hedger ◽

J. A. Muir ◽

W. R. Winnall

Keyword(s):

Sertoli Cell ◽

Inflammatory Cytokines ◽

Cell Function ◽

Inhibitory Effect ◽

Inhibitory Effects ◽

Tight Junction Formation ◽

Interleukin 1Α ◽

Factor Α ◽

Necrosis Factor

There is increasing evidence that the Sertoli cell, in addition to modulating responses to direct antigenic challenges (eg. intratesticular allografts), is central to the response of the testis to inflammation and infection. Systemic inflammation exerts an inhibitory effect on spermatogenesis, which has been attributed to the effects of fever, vascular disturbances, or loss of androgenic support. However, recent studies point to more direct effects of inflammation on spermatogenesis. The discovery that Sertoli cells express Toll-like receptors (TLR), and react to TLR ligands by producing inflammatory cytokines and other mediators, provides a mechanism to account for this direct inhibition. Moreover, the pattern of cytokines produced by the Sertoli cell during inflammation is highly characteristic. For example, when stimulated with TLR ligands the Sertoli cell produces the pro-inflammatory cytokines, interleukin-1α (IL1α) and IL6, and the regulatory cytokine, activin A, but does not produce IL1β and tumour necrosis factor-α, which are major pro-inflammatory products of activated macrophages. The disruptive effects of inflammation on spermatogenesis may be attributed to the elevated production of these cytokines, all of which have stimulatory or inhibitory effects on germ cell mitosis, meiosis and apoptosis and Sertoli cell tight junction formation, In addition, activation of TLR/IL1 mediated inflammatory pathways in the Sertoli cell inhibits its ability to respond to its principal trophic hormone, follicle-stimulating hormone. Studies on the regulation of these interactions will further establish the role of the Sertoli cell in inflammation and infection. However, such studies also have important implications for normal Sertoli cell function, as TLRs can respond to endogenous ligands as well. Consequently, the Sertoli cell may be viewed as a sentinel cell, supporting and protecting spermatogenesis when conditions are optimal, but rapidly shutting down spermatogenesis in the presence of infection or illness. Intriguingly, these apparently disparate roles appear to involve common inflammation-related mechanisms.

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Sodium iron EDTA and ascorbic acid, but not polyphenol oxidase treatment, counteract the strong inhibitory effect of polyphenols from brown sorghum on the absorption of fortification iron in young women

British Journal Of Nutrition ◽

10.1017/s0007114513002705 ◽

2013 ◽

Vol 111 (3) ◽

pp. 481-489 ◽

Cited By ~ 22

Author(s):

Colin I. Cercamondi ◽

Ines M. Egli ◽

Christophe Zeder ◽

Richard F. Hurrell

Keyword(s):

Ascorbic Acid ◽

Polyphenol Oxidase ◽

Young Women ◽

Inhibitory Effect ◽

Inhibitory Effects ◽

Fe Isotopes ◽

Absorption Studies ◽

Oxidase Enzyme ◽

Strong Inhibitory Effect

In addition to phytate, polyphenols (PP) might contribute to low Fe bioavailability from sorghum-based foods. To investigate the inhibitory effects of sorghum PP on Fe absorption and the potential enhancing effects of ascorbic acid (AA), NaFeEDTA and the PP oxidase enzyme laccase, we carried out three Fe absorption studies in fifty young women consuming dephytinised Fe-fortified test meals based on white and brown sorghum varieties with different PP concentrations. Fe absorption was measured as the incorporation of stable Fe isotopes into erythrocytes. In study 1, Fe absorption from meals with 17 mg PP (8·5 %) was higher than that from meals with 73 mg PP (3·2 %) and 167 mg PP (2·7 %;P< 0·001). Fe absorption from meals containing 73 and 167 mg PP did not differ (P= 0·9). In study 2, Fe absorption from NaFeEDTA-fortified meals (167 mg PP) was higher than that from the same meals fortified with FeSO4(4·6v.2·7 %;P< 0·001), but still it was lower than that from FeSO4-fortified meals with 17 mg PP (10·7 %;P< 0·001). In study 3, laccase treatment decreased the levels of PP from 167 to 42 mg, but it did not improve absorption compared with that from meals with 167 mg PP (4·8v.4·6 %;P= 0·4), whereas adding AA increased absorption to 13·6 % (P< 0·001). These findings suggest that PP from brown sorghum contribute to low Fe bioavailability from sorghum foods and that AA and, to a lesser extent, NaFeEDTA, but not laccase, have the potential to overcome the inhibitory effect of PP and improve Fe absorption from sorghum foods.

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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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A Critical Role of Nuclear m6A Reader YTHDC1 in Leukemogenesis by Regulating MCM Complex-Mediated DNA Replication

Blood ◽

10.1182/blood.2021011707 ◽

2021 ◽

Author(s):

Yue Sheng ◽

Jiangbo Wei ◽

Fang Yu ◽

Huanzhou Xu ◽

Chunjie Yu ◽

...

Keyword(s):

Dna Replication ◽

Critical Role ◽

Inhibitory Effect ◽

Self Renewal ◽

Hematopoietic Stem ◽

Strong Inhibitory Effect ◽

Mcm Complex ◽

Genetic Deletion

YTHDC1 has distinct functions as a nuclear N6-methyladenosine (m6A) reader in regulating RNA metabolism. Here we show that YTHDC1 is overexpressed in Acute Myeloid Leukemia (AML) and that it is required for proliferation and survival of human AML cells. Genetic deletion of Ythdc1 markedly blocks AML development and maintenance as well as self-renewal of leukemia stem cells (LSCs) in vivo in mice. We find that Ythdc1 is also required for normal hematopoiesis and hematopoietic stem/progenitor cell (HSPC) maintenance in vivo. Notably, Ythdc1 haploinsufficiency reduces self-renewal of LSCs, but not HSPCs in vivo. YTHDC1 knockdown has a strong inhibitory effect on proliferation of primary AML cells. Mechanistically, YTHDC1 regulates leukemogenesis through MCM4, which is a critical regulator of DNA replication. Our study provides the compelling evidence to show an oncogenic role and a distinct mechanism of YTHDC1 in AML.

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Inhibitory effects of calcium ionophore pretreatment of porcine oocytes on polyspermic fertilization

Zygote ◽

10.1017/s0967199406003558 ◽

2006 ◽

Vol 14 (1) ◽

pp. 17-22 ◽

Cited By ~ 2

Author(s):

T. Hayashi ◽

H. Sato ◽

H. Iwata ◽

T. Kuwayama ◽

Y. Monji

Keyword(s):

Calcium Ionophore ◽

Inhibitory Effect ◽

Calcium Ionophore A23187 ◽

The Other ◽

Ionophore A23187 ◽

Inhibitory Effects ◽

High Concentration ◽

Maturation Period ◽

Low Concentration ◽

Other Hand

The present study examined the inhibitory effects of various pretreatment concentrations (0–100 μM) of the calcium ionophore A23187 on polyspermic fertilization and then examined the effect of the maturation period and the time between calcium ionophore treatment and fertilization on the inhibitory effect of calcium ionophore on polyspermic fertilization. In experiment 1, a high concentration of calcium ionophore (100 μM) increased the rate of activated oocytes, but the rate of fertilization declined. On the other hand, when oocytes were treated with a low concentration of calcium ionophore (10 μM), monospermic fertilization was significantly increased (10 μM; 31.3%) (p < 0.05). In experiment 2, oocytes were cultured for various times (0, 0.5, 3, 6 h) after calcium ionophore treatment (10 μM) before fertilization. The highest rate of monospermic fertilization was detected in the oocytes cultured for 6 h after calcium ionophore treatment before fertilization. In experiments 3 and 4, we examined the effect of the maturation period (40 h or 44 h) on the rate of fertilization and blastulation of oocytes pretreated with calcium ionophore. The treatment of oocytes with calcium ionophore significantly decreased the rate of polyspermic fertilization regardless of the maturation period (44 h: with calcium ionophore 26.25% vs without 78.8%; 40 h: with calcium ionophore 37.5% vs without 77.5%); however, calcium ionophore treatment increased the rates of monospermic fertilization and blastulation of the oocytes matured for 44 h, but not those matured for 40 h. In conclusion, activation with a low concentration of calcium ionophore (10 μM) and a further 6 h of culture before fertilization improved the rate of monospermic fertilization and blastulation.

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Effects of periodontal dressings on fibroblasts and gingival wound healing in dogs

Acta Veterinaria Hungarica ◽

10.1556/avet.52.2004.1.4 ◽

2004 ◽

Vol 52 (1) ◽

pp. 33-46 ◽

Cited By ~ 4

Author(s):

M. Petelin ◽

Z. Pavlica ◽

Urška Batista ◽

Draga Štiblar-Martinčič ◽

U. Skalerič

Keyword(s):

Cell Proliferation ◽

Wound Healing ◽

Inflammatory Reaction ◽

Inhibitory Effect ◽

The Other ◽

Gingival Tissue ◽

Histological Evaluation ◽

Beagle Dogs ◽

Inhibitory Effects

In the present study the effects of different commercially available periodontal dressings (Peripac, Barricaid, Fittydent, Reso-Pack and Myzotect-tincture) on fibroblast (V-79-379A) proliferation and survival were tested in vitro. Barricaid, Fittydent and Reso-Pack periodontal dressings have only small inhibitory effects on cell proliferation (83.3 ± 9%, 71.6 ± 8.7% and 87.3 ± 4.5% of control after 48 h, respectively) in comparison with the great inhibitory effect of Myzotect-tincture (2.9 ± 0.1%) and Peripac (33.7 ± 11.4%) (p < 0.001). Barricaid was the only dressing where 41% of cells survived after exposure, while the other four dressings killed all the cells in 6 days. In addition, the healing of artificially created gingival wounds covered by Barricaid and Reso-Pack was followed for 7 days in 12 Beagle dogs. Histological evaluation of gingival tissue demonstrated that wounds covered by Reso-Pack showed the best epithelisation and vascularity and the least inflammatory reaction in first 4days. Later the observed parameters were similar with those of wounds covered by Barricaid or without pack. The present results indicate that Peripac periodontal dressing and Myzotect-tincture showed the highest cytotoxicity to fibroblasts in vitro. From the histological observations in Beagle dogs Reso-Pack has been found to be the most suitable dressing, followed by Barricaid.

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Role of cyclic GMP in the control of capacitative Ca2+ entry in rat pancreatic acinar cells

Biochemical Journal ◽

10.1042/bj3110649 ◽

1995 ◽

Vol 311 (2) ◽

pp. 649-656 ◽

Cited By ~ 27

Author(s):

P Gilon ◽

J F Obie ◽

X Bian ◽

G S J Bird ◽

J W Putney

Keyword(s):

Sodium Nitroprusside ◽

Cyclic Gmp ◽

Acinar Cells ◽

Inhibitory Effect ◽

Pancreatic Acinar Cells ◽

Inhibitory Effects ◽

Intracellular Cgmp ◽

Agonist Stimulation ◽

Ly 83583

We have investigated the possible roles of cyclic GMP (cGMP) in initiating or regulating capacitiative Ca2+ entry in rat pancreatic acinar cells. In medium containing 1.8 mM external Ca2+, thapsigargin activated Ca2+ entry and slightly but significantly increased intracellular cGMP concentration. This rise in cGMP levels was prevented by pretreating the cells with the guanylate cyclase inhibitor, LY-83583, or by omitting Ca2+ during stimulation by thapsigargin or methacholine. LY-83583 and NG-nitro-L-arginine (L-NA, an inhibitor of NO synthase) both had a small inhibitory effect on Ca2+ entry when they were added after thapsigargin in Ca2(+)-containing medium, and they reduced by 32 and 48% respectively the thapsigargin-induced capacitative Ca2+ entry when added to the cells during a 20 min preincubation period. However, neither dibutyryl cGMP (Bt2cGMP) nor sodium nitroprusside, an NO mimic, affected either basal intracellular Ca2+ concentration [Ca2+]i or thapsigargin-induced capacitative Ca2+ entry. Further, the inhibitory effects observed after preincubation with LY-83583 or L-NA could not be prevented by preincubation with Bt2cGMP, nor could they be reversed by adding Bt2cGMP, 8-bromo-cGMP or sodium nitroprusside acutely after activation of capacitative Ca2+ entry by thapsigargin. Finally, pretreatment of cells with LY-83583 or L-NA did not affect Ca2+ signalling in response to 1 microM methacholine, including the pattern of [Ca2+]i oscillations. In conclusion, in pancreatic acinar cells, the rise in cellular cGMP levels appears to depend on, rather than cause, the increase in [Ca2+]i with agonist stimulation.

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THE INFLUENCE OF CORTISONE ON EXPERIMENTAL VIRAL INFECTION

Journal of Experimental Medicine ◽

10.1084/jem.106.6.883 ◽

1957 ◽

Vol 106 (6) ◽

pp. 883-892 ◽

Cited By ~ 7

Author(s):

Edwin D. Kilbourne ◽

Keyword(s):

Protein Synthesis ◽

Influenza Virus ◽

Potent Inhibitor ◽

Inhibitory Effect ◽

The Self ◽

Inhibitory Effects ◽

Obligate Intracellular ◽

Intracellular Parasites ◽

Experimental Viral Infection

Cortisone is a highly potent inhibitor of influenza virus synthesis in the chick embryo, inducing manifest inhibition in doses of 0.1 to 1.0 µg/egg. Inhibition of viral synthesis is only temporarily manifest. As infection continues, the negation by cortisone of the self-limiting effects of viral autointerference obscures the coincident cortisone effect on synthesis. The inhibitory effect of cortisone may be induced late in the course of viral multiplication, after conclusion of the latent period. It is proposed that inhibition of viral synthesis with cortisone is a corollary of the steroid's inhibitory effects on growth and protein synthesis of the infected host. The role of adrenal corticoids in the regulation of infection with obligate intracellular parasites deserves continued investigation.

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Effect of calmodulin on the stimulation of capacitation and acrosome reaction of frozen thawed bull spermatozoa

Bangladesh Journal of Animal Science ◽

10.3329/bjas.v45i3.31033 ◽

2017 ◽

Vol 45 (3) ◽

pp. 1-9

Author(s):

QS Akter ◽

KMA Tareq ◽

K Hamano ◽

RB Gilchrist

Keyword(s):

Acrosome Reaction ◽

Specific Binding ◽

Staining Technique ◽

Inhibitory Effect ◽

Free Medium ◽

Inhibitory Effects ◽

Specific Binding Protein ◽

Bovine Spermatozoa ◽

Mammalian Spermatozoa

Capacitation and acrosome reaction (AR) are the prerequisites for successful fertilization by mammalian spermatozoa. Intracellular calcium (Ca2+) has a regulatory role in sperm motility, capacitation, and AR. Calmodulin (CaM) antagonists calmidazolium (CZ) and N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W7) were used to investigate the possible role of CaM, a Ca2+ specific binding protein, on motility, capacitation and AR of frozen-thawed bovine spermatozoa. Capacitation and AR in sperm were evaluated by using chlortetracycline (CTC) staining technique. Addition of the 1 mM dibutyryl cAMP (dbcAMP) and 100 ?M 1-methy l-3-isobutylxanthine (IBMX) to CaM antagonists treated sperm incubated in the presence of NaHCO3 and CaCl2 in media overcome the inhibitory effects of these antagonists to support capacitation and AR at 4 h of incubation period. In contrast, addition of dbcAMP with IBMX induced AR in spermatozoa incubated with NaHCO3-free medium but these compounds did not induce AR in cells incubated in CaCl2-free medium. However, the addition of dbcAMP and IBMX partially, but significantly (p<0.01) reversed the inhibitory effect of W7 and CZ on the sperm capacitation and AR. These results suggest that CaM may play an important role in the regulation of capacitation and AR in frozen-thawed bovine spermatozoa.Bang. J. Anim. Sci. 2016. 45 (3): 1-9

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