scholarly journals Mechanical stimulation induces formin-dependent assembly of a perinuclear actin rim

2015 ◽  
Vol 112 (20) ◽  
pp. E2595-E2601 ◽  
Author(s):  
Xiaowei Shao ◽  
Qingsen Li ◽  
Alex Mogilner ◽  
Alexander D. Bershadsky ◽  
G. V. Shivashankar
Keyword(s):  
Actin Polymerization ◽  
Genetic Diseases ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Actin Dynamics ◽  
Ionophore A23187 ◽  
Dependent Manner ◽  
Mechanical Stimuli ◽  
Actin Assembly ◽  
Force Application

Cells constantly sense and respond to mechanical signals by reorganizing their actin cytoskeleton. Although a number of studies have explored the effects of mechanical stimuli on actin dynamics, the immediate response of actin after force application has not been studied. We designed a method to monitor the spatiotemporal reorganization of actin after cell stimulation by local force application. We found that force could induce transient actin accumulation in the perinuclear region within ∼2 min. This actin reorganization was triggered by an intracellular Ca2+ burst induced by force application. Treatment with the calcium ionophore A23187 recapitulated the force-induced perinuclear actin remodeling. Blocking of actin polymerization abolished this process. Overexpression of Klarsicht, ANC-1, Syne Homology (KASH) domain to displace nesprins from the nuclear envelope did not abolish Ca2+-dependent perinuclear actin assembly. However, the endoplasmic reticulum- and nuclear membrane-associated inverted formin-2 (INF2), a potent actin polymerization activator (mutations of which are associated with several genetic diseases), was found to be important for perinuclear actin assembly. The perinuclear actin rim structure colocalized with INF2 on stimulation, and INF2 depletion resulted in attenuation of the rim formation. Our study suggests that cells can respond rapidly to external force by remodeling perinuclear actin in a unique Ca2+- and INF2-dependent manner.

Immunologic mast cell-mediated responses and histamine release are attenuated by heparin

1992 ◽  
Vol 73 (3) ◽  
pp. 1093-1101 ◽  
Author(s):  
J. Lucio ◽  
J. D'Brot ◽  
C. B. Guo ◽  
W. M. Abraham ◽  
L. M. Lichtenstein ◽  
...  
Keyword(s):  
Mast Cell ◽  
Mast Cells ◽  
Histamine Release ◽  
Immunoglobulin E ◽  
Specific Antigen ◽  
Calcium Ionophore ◽  
Intradermal Injection ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Dependent Manner

Heparin has been shown to act as a competitive inhibitor of inositol 1,4,5-triphosphate (InsP3) receptors in various cell types. Because InsP3 is one of the second messengers involved in stimulus-secretion coupling in mast cells, it is possible that heparin may inhibit mast cell-mediated reactions. Therefore, in allergic sheep, we tested this hypothesis in two mast cell-mediated reactions induced by immunologic and nonimmunologic stimuli: immediate cutaneous reaction (ICR) and acute bronchoconstrictor response (ABR). In 12 sheep allergic to Ascaris suum antigen, the surface area of the skin wheal was determined 20 min after intradermal injection (0.05 ml) of increasing concentrations of specific antigen, compound 48/80, and histamine, without and after pretreatment with heparin (100, 300, or 1,000 U/kg i.v.). Antigen, compound 48/80, and histamine produced concentration-dependent increases in ICR. Heparin “partially” inhibited the ICR to antigen and compound 48/80 in a dose-dependent manner without modifying the ICR to histamine. The heparin preservative benzyl alcohol was ineffective. In 11 additional sheep, specific lung resistance was measured before and after inhalation challenges with antigen, compound 48/80, and histamine without and with aerosol heparin pretreatment (1,000 U/kg). Heparin blocked the antigen- and compound 48/80-induced bronchoconstriction without modifying the airway effects of histamine. In isolated human uterine mast cells, heparin inhibited the anti-immunoglobulin E- but not the calcium ionophore- (A23187) induced histamine release. These data suggest that heparin inhibits the ICR and ABR induced by stimuli that produce immunologic and nonimmunologic mast cell degranulation without attenuating the effects of histamine.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Platelet-activating factor both stimulates and "primes" human polymorphonuclear leukocyte actin filament assembly

Blood ◽  
1987 ◽  
Vol 70 (6) ◽  
pp. 1921-1927 ◽  
Author(s):  
M Shalit ◽  
GA Dabiri ◽  
FS Southwick
Keyword(s):  
Actin Filament ◽  
Polymorphonuclear Leukocyte ◽  
Actin Polymerization ◽  
Platelet Activating Factor ◽  
Calcium Ionophore ◽  
Leukotriene B4 ◽  
Calcium Ionophore A23187 ◽  
Optimal Concentration ◽  
Ionophore A23187 ◽  
Filament Assembly

Abstract The phospholipid inflammatory mediator, platelet-activating factor (PAF), can stimulate polymorphonuclear leukocyte (PMN) chemotaxis. Conversion of cytoplasmic actin from monomers to filaments is associated with PMN motile functions. Using the fluorescent actin filament stain nitrobenzodiaxole phallicidin, we have investigated PAF's effects on human PMN actin polymerization. Concentrations of PAF between 1 x 10(-11) to 1 x 10(-6) mol/L induced actin filament (F- actin) assembly. An optimal concentration of PAF (1–5 x 10(-8) mol/L) induced a significantly lower rise in relative F-actin content (1.72 +/- 0.07 SEM) than an optimal concentration (5 x 10(-7) mol/L) of the chemotactic peptide FMLP (2.21 +/- 0.06). Unlike FMLP (F-actin content: 1.25 +/- 0.04 at five seconds), PAF stimulation was associated with a delay of more than five seconds (1.04 +/- 0.01 at five seconds) before an increase in F-actin could be detected. F-actin concentration reached maximum levels by 30 to 60 seconds. Prolonged stimulation (20 minutes) with PAF was associated with two phases of polymerization and depolymerization. Like FMLP, the initiation of actin filament assembly by PAF required receptor occupancy, this reaction being totally blocked by the PAF receptor inhibitor, SKI 63–441. As evidenced by the lack of inhibition by nordihydroguaiaretic acid (5 to 20 mumol/L), the production of leukotriene B4 was not required for the PAF-induced changes in F-actin. Like FMLP, PAF's ability to stimulate PMN actin polymerization was inhibited by pertussis toxin (.05 to 2.5 micrograms/mL) but not impaired by the addition of EGTA and/or the calcium ionophore A23187. Preincubation with 1 x 10(-11) to 1 x 10(-8) mol/L PAF for 2 to 60 minutes enhanced the rise in F-actin content induced by low concentrations of FMLP (5 x 10(-12) to 1 x 10(-10) mol/L) indicating that this phospholipid was capable of “priming” the PMN actin polymerization response.

Ca2+-dependent Activation of the 33-kDa Protein Kinase Transmits Thrombin Receptor Signals in Human Platelets

1996 ◽  
Vol 76 (03) ◽  
pp. 439-443 ◽  
Author(s):  
Masaru Ido ◽  
Shinya Kato ◽  
Hiroyuki Ogawa ◽  
Kenji Hayashi ◽  
Yoshihiro Komada ◽  
...  
Keyword(s):  
Kinase Inhibitor ◽  
Calcium Ionophore ◽  
Structural Analog ◽  
Human Platelets ◽  
Calcium Ionophore A23187 ◽  
Similar Degree ◽  
Ionophore A23187 ◽  
Dependent Manner ◽  
Serine Threonine Kinase ◽  
Acetoxymethyl Ester

SummaryThrombin stimulation induces a dramatic increase in the activity of the 33-kDa serine/threonine kinase (PK33) in human platelets (10). The Arg-Gly-Asp (RGD) peptide, an inhibitor of the thrombin-mediated aggregation of platelets, did not affect the PK33 activation induced by thrombin suggesting that the activation of this kinase occurs independently from platelet aggregation. To identify a potential role of Ca2+ and calmodulin in the regulation of PK33, the effect of several Ca2+/calmodulin inhibitors on the thrombin-induced activation of PK33 was assessed using denaturation/renaturation method. Pretreatment of platelets with EGTA decreased the maximum PK33 activity induced by thrombin. The chelation of both the extra- and the intracellular Ca2+ by EGTA and by acetoxymethyl ester of 5,5′ -dimethyl-bis-(<9-aminophen-oxy) ethane-N,N,N′,N′-tetraacetic acid (BAPTA-AM) decreased further the PK33 activation by thrombin. Preincubation of platelets with the anticalmodulin agent, N-(4-aminobutyl)-5-chloro-2-naphtha-lenesulfonamide (W13), inhibited markedly the activation of PK33 by thrombin, whereas the inactive structural analog N-(4-aminobutyl)-2-naphthalenesulfonamide (W12) and the myosin light chain kinase inhibitor 1 -(5-chloronaphthalene-1 -sulfonyl)-1 H-hexahydro-1,4-diaze-pine (ML9) showed very weak inhibitory effects. Treatment of resting platelets with the calcium ionophore, A23187, activated PK33 in a dose-dependent manner; phorbol 12-myristate 13-acetate enhanced this effect. However, the two foregoing agents did not induce similar degree of PK33 activities as thrombin. These results indicate that the activation of PK33 is independent of the formation of the GPIIb/IIIa-fibrinogen complex and that it might be regulated by a Ca2+-dependent pathway.

scholarly journals The Role of Rhodomyrtus tomentosa (Aiton) Hassk. Fruits in Downregulation of Mast Cells-Mediated Allergic Responses

2019 ◽  
Vol 2019 ◽  
pp. 1-7 ◽  
Author(s):  
Thanh Sang Vo ◽  
Young-Sang Kim ◽  
Dai-Nghiep Ngo ◽  
Dai-Hung Ngo
Keyword(s):  
Mast Cells ◽  
Biological Activities ◽  
Interleukin 1 ◽  
Reactive Oxygen Species Generation ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Flowering Plant ◽  
Ionophore A23187 ◽  
Dependent Manner ◽  
Rhodomyrtus Tomentosa

Rhodomyrtus tomentosa, a flowering plant of Myrtaceae family from southern and southeastern Asia, was known to possess a rich source of structurally diverse and various biological activities. In this study, the inhibitory effect of R. tomentosa fruit extract (RFE) on allergic responses in calcium ionophore A23187-activated RBL-2H3 mast cells was investigated. The result showed that RFE was able to inhibit mast cell degranulation via decreasing β-hexosaminidase release and intracellular Ca2+ elevation at the concentration of 400 μg/ml. Moreover, the suppressive effects of RFE on the production of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) were evidenced. In addition, RFE effectively scavenged DPPH radical and suppressed the reactive oxygen species generation in a dose-dependent manner. Notably, the pretreatment of RFE caused the downregulation of tyrosine kinase Fyn phospholipid enzyme phospholipase Cγ (PLCγ), extracellular-signal-regulated kinase (ERK), and nuclear factor kappa B (NF-κB) phosphorylation. These results indicated that RFE could be a promising inhibitor of allergic responses and may be developed as bioactive ingredient for prevention or treatment of allergic diseases.

Canine jejunal submucosa cultures: characterization and release of neural somatostatin

1990 ◽  
Vol 68 (6) ◽  
pp. 705-710 ◽  
Author(s):  
A. M. J. Buchan ◽  
A. D. Doyle ◽  
E. Accili
Keyword(s):  
Substance P ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Neuronal Cultures ◽  
Ionophore A23187 ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Cell Content ◽  
Cell Extracts

A primary culture of the canine jejunal submucosa has been established and used to investigate neuronal somatostatin release. Immunocytochemical characterization of the cultures demonstrated the presence of the following peptidergic neurons: neurotensin (30%), somatostatin (27%), vasoactive intestinal polypeptide (14%), neuropeptide Y (10%), and substance P (5%). No immunoreactive neurons were observed with the available antisera to galanin, gastrin-releasing peptide, and motilin. The concentration of somatostatin-like immunoreactivity, as determined by radioimmunoassay of cell extracts, was 358 ± 105 pmol/well. Basal release of somatostatin was 4.4 ± 0.9% total cell content and was significantly inhibited by the addition of substance P at 1 and 100 nM. The addition of the calcium ionophore, A23187, with phorbol 12-myristate 13-acetate stimulated somatostatin release in a concentration-dependent manner. These data indicate that short-term cultures of the jejunal submucosal plexus will be an excellent model for determination of the factors influencing the release of neural somatostatin.Key words: immunocytochemistry, neuronal cultures, neurofilament, substance P.

Inhibition of Human Platelet Functions by Verapamil

1981 ◽  
Vol 45 (02) ◽  
pp. 158-161 ◽  
Author(s):  
Y Ikeda ◽  
M Kikuchi ◽  
K Toyama ◽  
K Watanabe ◽  
Y Ando
Keyword(s):  
Platelet Aggregation ◽  
Calcium Ionophore ◽  
Serotonin Release ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Dependent Inhibition ◽  
Platelet Functions

SummaryThe effects of verapamil, a coronary vasodilator, on platelet functions were studied.Platelet aggregation induced by ADP, epinephrine or collagen was inhibited by verapamil in vitro. Calcium ionophore A23187-induced platelet aggregation was also inhibited by verapamil in a concentration dependent manner. In washed platelets, verapamil caused a dose-dependent inhibition of serotonin release induced either by thrombin or A23187 in the absence of extracellular calcium. Addition of 1 mM CaCl2 with A23187 or thrombin partially overcame this inhibition. Addition of 1 mM CaCl2 in the absence of verapamil had no effect on thrombin- or A23187-induced secretion. When verapamil was administered to the healthy volunteers at the dosage commonly used, inhibition of platelet aggregation was observed 2 hrs after the drug ingestion. It is of great interest that verapamil potentiated the anti-aggregating activity of prostacyclin in vitro.Our results may suggest a potential role for verapamil in the treatment of thrombotic disorders.

Delayed activation of nociceptors: correlation with delayed pain sensations induced by sustained stimuli

1991 ◽  
Vol 66 (3) ◽  
pp. 729-734 ◽  
Author(s):  
D. M. White ◽  
Y. O. Taiwo ◽  
T. J. Coderre ◽  
J. D. Levine
Keyword(s):  
Calcium Ionophore ◽  
Intradermal Injection ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Mechanical Stimuli ◽  
Mechanical Threshold ◽  
Electrophysiological Studies ◽  
Normal Rat ◽  
Short Time ◽  
Withdrawal Response

1. In this study we used psychophysical experiments in humans and behavioral and electrophysiological studies in rats to evaluate nociceptive and C-fiber mechanoheat nociceptor (C-MH) responses to sustained mechanical stimuli that are initially nonpainful or nonnoxious. 2. In normal rat skin, sustained subthreshold mechanical stimuli activate C-MHs (n = 36) with a delayed onset that parallels the delayed pain sensation recorded in human psychophysical tests. 3. The subthreshold stimuli did not induce a decrease in mechanical threshold (n = 11), and the effect of the subthreshold stimulus on latency to firing of C-MHs (n = 6) persists for a very short time after the stimulus is removed (less than 10 s). 4. Intradermal injection of prostaglandin E2 (PGE2; 100 ng), which induced a significant decrease in the mechanical threshold of C-MHs (n = 7), had no effect on the latency of the delayed activation of C-MHs. Also, indomethacin, which inhibits the synthesis of prostaglandins, had no effect on the latency of the delayed paw-withdrawal response in the behavioral test. 5. Intradermal injection of the calcium ionophore A23187 significantly reduced the latency of the delayed activation of C-MHs (n = 6) the calcium chelator Quin 2 (n = 6) significantly increased the latency. A23187 and Quin 2 had similar effects on the latencies to paw withdrawal in behavioral tests. The sensitization of C-MHs (n = 9) by PGE2 was not, however, affected by Quin 2.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Increased expression of procoagulant activity on the surface of human platelets exposed to heavy-metal compounds

1995 ◽  
Vol 308 (1) ◽  
pp. 15-21 ◽  
Author(s):  
C A Goodwin ◽  
C P D Wheeler-Jones ◽  
S Namiranian ◽  
S Bokkala ◽  
V V Kakkar ◽  
...  
Keyword(s):  
Heavy Metal ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Procoagulant Activity ◽  
Ionophore A23187 ◽  
Dependent Manner ◽  
Anionic Phospholipid ◽  
Metal Compounds ◽  
Platelet Surface ◽  
Thrombin Formation

One of the essential roles for platelets in haemostasis is in the potentiation of blood clotting due to the contribution of anionic phospholipid from the surface of the cells, as an essential cofactor to the proteolytic reactions of coagulation (platelet procoagulant activity). Only a limited number of agonists are known to initiate platelet procoagulant activity. In this study the rate of thrombin formation on the platelet surface was observed to increase in a dose-dependent manner upon treatment of washed platelets with heavy-metal compounds. Unlike the immediate increase observed upon treatment of platelets with calcium ionophore, A23187, the change due to these agents was progressive, approaching a maximum after 10 min. The maximum-fold acceleration of the rate of thrombin formation compared with control platelets was calculated for HgCl2 (56-fold), AgNO3 (42-fold) phenylmercuriacetate (24-fold) and thimerosal (14-fold), compared with 70-fold observed for calcium ionophore. The increase in procoagulant activity due to HgCl2 coincided with a large increase in intracellular calcium and phosphorylation of 22 and 45 kDa proteins. It is considered that the mechanism responsible for the increase in procoagulant activity is exposure of anionic phospholipids. This was detected by a 2-fold increase in the binding of 125I-annexin V upon addition of HgCl2, compared with resting platelets (3-fold on treatment of platelets with calcium ionophore). In contrast to the generation of activity by A23187 and other known agonists of this reaction, heavy-metal compounds appeared to cause little or no release of microparticles from the platelet surface. Since HgCl2 did not cause aggregation of platelets or significant release of serotinin, these findings may give further support to the need for exposure and ligation of glycoprotein IIb:IIIa for vesiculization to occur. Treatment of platelets with heavy metals may constitute a new approach to investigating the early changes in the cell membrane which lead to increased expression of anionic phospholipid.

Effect of the calcium ionophore A23187 on superoxide generation in phorbol ester stimulated human neutrophils

1985 ◽  
Vol 63 (12) ◽  
pp. 1543-1546 ◽  
Author(s):  
Colette F. Strnad ◽  
Kenneth Wong
Keyword(s):  
Protein Kinase C ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Superoxide Production ◽  
Human Neutrophils ◽  
Ionophore A23187 ◽  
Dependent Manner ◽  
Superoxide Generation ◽  
Kinase C ◽  
Dose Dependent

The calcium ionophore, A23187, and the tumor-promoting phorbol ester, phorbol 12-myristate 13-acetate (PMA), interacted synergistically to elicit an accelerated superoxide production response in human neutrophils. The lag period preceding PMA-induced superoxide generation was decreased in a dose-dependent manner by A23187 at a concentration range from 1.0 × 10−8 to 1.0 × 10−5 M. Superoxide production rate, however, was subject to biphasic effects. While the rate was potentiated in a dose-dependent manner at A23187 concentrations below 1.0 × 10−6 M, inhibitory influences became manifest at higher concentrations. Total superoxide production was subject to inhibitory effects, characterized by a mean inhibitory dose of 1.3 × 10−6 M. The synergistic interaction of A23187 with PMA is consistent with a role for protein kinase C in neutrophil activation. Inhibition at high A23187 concentrations appeared to result from the effects of elevated intracellular Ca2+ levels on either NADPH oxidase itself, or some step in the transduction process linking protein kinase C to the oxidase complex.

scholarly journals Trace levels of peptidoglycan in serum underlie the NOD-dependent cytokine response to endoplasmic reticulum stress

2019 ◽  
Author(s):  
Raphael Molinaro ◽  
Robert Flick ◽  
Dana J. Philpott ◽  
Stephen E. Girardin
Keyword(s):  
Cell Culture ◽  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Calf Serum ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Dependent Manner ◽  
Intracellular Ca ◽  
Inflammatory Signalling

AbstractNOD1 and NOD2 are intracellular sensors of bacterial peptidoglycan that belong to the Nod-like receptor (NLR) family of innate immune proteins. In addition to their role as direct bacterial sensors, it was recently proposed that NOD proteins could detect endoplasmic reticulum (ER) stress induced by thapsigargin, an inhibitor of the sarcoplasmic or endoplasmic reticulum calcium ATPase family (SERCA) that pumps Ca2+into the ER, resulting in pro-inflammatory signalling. Here, we confirm that thapsigargin induces pro-inflammatory signalling in epithelial cells in a NOD-dependent manner. However, the effect was specific to thapsigargin, as tunicamycin and the subtilase cytotoxin SubAB from Shiga toxigenicEscherichia coli,which induce ER stress by other mechanisms, did not induce cytokine expression. The calcium ionophore A23187 also induced NOD-dependent signalling, and the calcium chelator BAPTA-AM blunted thapsigargin-dependent pro-inflammatory signalling, showing NOD proteins responded to a rise in intracellular Ca2+. Since intracellular Ca2+directly affects vesicular trafficking, we tested if thapsigargin-induced NOD activation required endocytosis. Our results demonstrate that both endocytosis and the addition of serum to the cell culture medium were required for thapsigargin-mediated NOD activation. Finally, we analyzed cell culture grade fetal calf serum as well as serum from laboratory mice by high-pressure liquid chromatography and mass spectrometry, and identified the presence of various peptidoglycan fragments. We propose that cellular perturbations that affect intracellular Ca2+can trigger internalization of peptidoglycan trace contaminants found in culture serum, thereby stimulating pro-inflammatory signalling. The presence of peptidoglycan in animal serum suggests that a homeostatic function of NOD signalling may have been previously overlooked.

Sign in / Sign up

Export Citation Format

Share Document