The kinetics of chemotactic peptide-induced change in F-actin content, F-actin distribution, and the shape of neutrophils.

Howard, Thomas H.; Oresajo, Christian

doi:10.1083/jcb.101.3.1078

Cited by 301 publications

(191 citation statements)

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“…Actin polymerization was evaluated by a flow cytometric assay, as described [16,17]. For each experiment, isolated neutrophils (10 5 ) were put in tubes identified as follows: (1) blank; (2) control sample with resting neutrophils stored at 48C before labeling with FITC-phalloidin; (3) control sample with neutrophils incubated at 378C for 120 s before labeling with FITC-phalloidin; (4) to (7) neutrophils incubated at 378C for 120 s and then stimulated with fMLP (10 À8 M final concentration), for 15, 30, 60, and 120 s, respectively.…”

Section: Flow Cytometric Evaluation Of Actin Polymerizationmentioning

confidence: 99%

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Actin polymerization in neutrophils from donors of peripheral blood stem cells: Divergent effects of glycosylated and nonglycosylated recombinant human granulocyte colony‐stimulating factor

et al. 2006

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Neutrophil functions can be modified by Recombinant human G-CSF (rhG-CSF) treatment, with divergent effects on phagocytosis, motility, bactericidal activity, and surface molecule expression. Neutrophil morphology is modified by treatment with filgrastim (the nonglycosylated form of rhG-CSF), while it is not affected by lenograstim (the glycosylated type of rhG-CSF). Little information is available about actin polymerization in neutrophils from subjects treated with the two types of rhG-CSF. In the current paper we evaluated two groups of donors of peripheral blood stem cells (PBSC) for allogeneic transplantation. Ten subjects were treated with filgrastim and 10 with lenograstim to mobilize PBSC; 15 blood donors were evaluated as a control group. Actin polymerization (both spontaneous and fMLP-stimulated) was studied by a flow cytometric assay. A microscopic fluorescent assay was also carried out to evaluate F-actin distribution in neutrophils. We found that filgrastim induced an increased F-actin content in resting neutrophils, along with morphologic evidence for increased actin polymerization distributed principally at the cell membrane and frequently polarized in focal areas; in addition, fMLP was not able to induce further actin polymerization. On the contrary, treatment with lenograstim was associated with F-actin content, distribution, and polymerization kinetics indistinguishable from those displayed by control neutrophils. Such experimental results show that filgrastim and lenograstim display divergent effects also on neutrophil actin polymerization and provide further explanation for previous experimental findings. Am. J. Hematol. 81:318-323, 2006. V V C 2006 Wiley-Liss, Inc.

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Section: Flow Cytometric Evaluation Of Actin Polymerizationmentioning

confidence: 99%

“…F-actin content was calculated as described [16,17]. The FL peak channel number of each histogram was recorded, and values of relative F-actin content were expressed as the ratio of the test peak channel number to the control peak channel number.…”

Section: Flow Cytometric Evaluation Of Actin Polymerizationmentioning

confidence: 99%

Actin polymerization in neutrophils from donors of peripheral blood stem cells: Divergent effects of glycosylated and nonglycosylated recombinant human granulocyte colony‐stimulating factor

et al. 2006

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“…42 During the steady crawling phase, the fraction of polymerized actin returns to near pre-stimulation levels. 11,22 It is sometimes assumed that this return is due to eventual recapping of filaments to their prestimulation levels, but our model results suggest that recapping is not necessary as the steady state polymer fraction is not a strong indicator of the fraction of free barbed ends.…”

Section: Signature Effectsmentioning

confidence: 71%

Relationships between Actin Regulatory Mechanisms and Measurable State Variables

Bindschadler

McGrath

2007

Ann Biomed Eng

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Abstract-In this report we extend our recent mathematical formulation of the actin cycle model [Bindschadler et al. Biophys. J. 86 (2004) 2720] to predict the influence of key regulatory mechanisms on network-scale state variables estimable in live cell experiments. Specifically, we examine the influence of regulation by cofilin, profilin, capping protein and proteins that adjust filament number through nucleation and/or filament severing, on the higher order variables of average filament length, polymer fraction, and filament turnover rate. Importantly, we find that severing/ nucleation, the acceleration of ADP-subunit disassembly by cofilin, and the catalytic and shuttle functions of profilin have Ôsignature' effects on the higher order state variables. In this way, measurement of the state variables in live cells can allow inference of regulatory mechanism(s) underlying changes in cell state. Our results compare favorably to published data for endothelial cells undergoing a transition from non-motile confluent cells to highly motile subconfluent cells. The extension of our model to higher order state variables allows us to investigate other important issues such as the distinction between basic and higher order measures of filament dynamics, the influence of thymosin b4 on network state variables, the interplay between thymosin b4 and profilin, and the synergystic effects of cofilin and profilin.

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“…The affinity constant of actin monomer (G-actin) for the barbed-end is 10-fold greater than that for the pointed end (Korn et al, 1987) and under identical buffer conditions the critical concentration for polymerization at the two ends differ dramatically. Because the barbed end is the preferred end for filament growth and the majority (-8 80%) of the ligand activated actin polymerization in nonmuscle cells like platelets and PMNs occurs on barbed-end nuclei (Rao and Varani, 1982;Fechheimer and Zigmond, 1983;Fox and Phillips, 1983;Howard and Oresajo, 1985b;Wallace et al, 1985;Lind et al, 1987;Howard et al, 1990a,b), the creation and regulation of barbed-end nuclei is a critical step in regulating actin polymerization in PMNs.…”

Section: Ga Interactions In Basal Etf-pmnsmentioning

confidence: 99%

“…Cells were analyzed on FACStar flow cytometer (Becton Dickinson, Mountain View, CA) as previously described (Howard and Oresajo, 1985b). The results are expressed as the relative F-actin content indicated by mean fluorescence channel number of test sample/mean fluorescence channel number of cells simultaneously exposed to FMLP and fixative at 0 s. The values presented are the means ± SD from multiple trials.…”

Section: Quantification Of F-actin Content By Nbdphallacidin Bindingmentioning

confidence: 99%

Role of gelsolin interaction with actin in regulation and creation of actin nuclei in chemotactic peptide activated polymorphonuclear neutrophils.

Deaton

Guerrero

Howard

1992

MBoC

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In vitro Ca++ activates gelsolin to sever F-actin and form a gelsolin-actin (GA) complex at the + end of F-actin that is not dissociated by ethylene glycol-bis(3-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) but is separated by EGTA + PIP/PIP2. The gelsolin blocks the + end on the actin filament, but the -end of the filament can still initiate actin polymerization. In thrombin activated platelets, evidence suggests that severing of F-actin by gelsolin increases GA complex, creates one -end actin nucleus and one cryptic + end actin nucleus per cut, and then dissociates to yield free + ends to nucleate rapid actin assembly. We examined the role of F-actin severing in creation and regulation of nuclei and polymerization in polymorphonuclear neutrophils (PMNs). At 2-s intervals after formyl peptide (FMLP) activation of endotoxin free (ETF) PMNs, change in GA complex was correlated with change in + end actin nuclei, -end actin nuclei, and F-actin content. GA complex was quantitated by electrophoretograms of proteins absorbed by antigelsolin from cells lysed in 10 mM EGTA, + end actin nuclei as cytochalasin (CD) sensitive and -end actin nuclei as CD insensitive increases in G-pyrenyl actin polymerization rates induced by the same PMNs, and F-actin content by NBDphallacidin binding to fixed cells. Thirty three percent of gelsolin was in GA complex in basal ETF PMNs; from 2-6 s, GA complexes dissociate (low = 15% at 10 s) and sequentially + end nuclei and F-actin content and then -end nuclei increase to a maximum at 10 s. At >40 s GA complex increase toward basal and + end nuclei and F-actin content returned toward basal. These kinetic data show gelsolin regulates availability of + end nuclei and actin polymerization in FMLP. However, absence of an initial increase in GA complex or -end nucleating activity shows FMLP activation does not cause gelsolin to sever F-or to bind G-actin to create cryptic + end nuclei in PMNs; the results suggest the + nucleus formation is gelsolin independent.

show abstract

The kinetics of chemotactic peptide-induced change in F-actin content, F-actin distribution, and the shape of neutrophils.

Cited by 301 publications

References 29 publications

Actin polymerization in neutrophils from donors of peripheral blood stem cells: Divergent effects of glycosylated and nonglycosylated recombinant human granulocyte colony‐stimulating factor

Actin polymerization in neutrophils from donors of peripheral blood stem cells: Divergent effects of glycosylated and nonglycosylated recombinant human granulocyte colony‐stimulating factor

Relationships between Actin Regulatory Mechanisms and Measurable State Variables

Role of gelsolin interaction with actin in regulation and creation of actin nuclei in chemotactic peptide activated polymorphonuclear neutrophils.

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