Effective neutrophil chemotaxis is strongly influenced by mean IL-8 concentration

Lin, Francis; Nguyen, C.; Wang, Shur-Jen; Saadi, Wajeeh; Gross, Steven P.; Jeon, Noo Li

doi:10.1016/j.bbrc.2004.05.029

Cited by 127 publications

(58 citation statements)

References 22 publications

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“…Such shallow channels are adequate for many microfluidic applications but not amenable for use with large mammalian cells (.10 mm in diameter) as well as other applications, such as flowing chemotactic gradients across adherent cells in a channel with minimal shearing. 5 While Lago et al 6 introduced a way to circumvent the height limitation of single-layer ink by printing up to four times using a thermal toner transfer method onto a glass substrate, the maximum height obtained with this approach was 25 mm. Vullev et al 7 demonstrated a non-lithographic fabrication approach of microfluidic devices by printing positive-relief masters with a laser-jet printer for detecting bacterial spores; the height of the channels, which is likewise dependent on the height of the ink, is limited to between 5 and 9 mm.…”

Section: Introductionmentioning

confidence: 99%

Shrinky-Dink microfluidics: rapid generation of deep and rounded patterns

et al. 2008

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We present a rapid and non-photolithographic approach to microfluidic pattern generation by leveraging the inherent shrinkage properties of biaxially oriented polystyrene thermoplastic sheets. This novel approach yields channels deep enough for mammalian cell assays, with demonstrated heights up to 80 mm. Moreover, we can consistently and easily achieve rounded channels, multi-height channels, and channels as thin as 65 mm in width. Finally, we demonstrate the utility of this simple microfabrication approach by fabricating a functional gradient generator. The whole process-from device design conception to working device-can be completed within minutes.

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Section: Introductionmentioning

confidence: 99%

Shrinky-Dink microfluidics: rapid generation of deep and rounded patterns

et al. 2008

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“…Generation of stable concentration profiles with linear or polynomial shapes has been enabled by recently developed microfluidic gradient-making networks (7)(8)(9)(10), and by the use of integrated microvalves (11), which allow such gradients to be imposed within a few seconds (12). Nevertheless, reports on chemotaxis in microfluidic devices have not addressed the issue of temporal vs. spatial sensing, whereas dependence of the efficiency of chemotaxis on attractant concentration, although discussed (9,13,14), has not been carefully studied.…”

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confidence: 99%

Bound attractant at the leading vs. the trailing edge determines chemotactic prowess

Herzmark¹,

Campbell

Wang³

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

106

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We have analyzed chemotaxis of neutrophil-differentiated HL60 cells in microfluidic devices that create exponential gradients of the chemoattractant, f-Met-Leu-Phe (fMLP). Such gradients expose each cell to a difference in fMLP concentration (⌬C) across its diameter that is directly proportional to the ambient concentration (C) at that cell's position in the gradient, so the ratio ⌬C/C is constant everywhere. Cells exposed to ambient fMLP concentrations near the constant of dissociation (K d) for fMLP binding to its receptor (Ϸ10 nM) crawl much less frequently when ⌬C/C is 0.05 than when it is 0.09 or 0.13. Hence, cells can detect the gradient across their diameter without moving and, thus, without experiencing temporal changes in attractant concentration. At all ⌬C/C ratios tested, the average chemotactic prowess of individual cells (indicated by the distance a cell traveled in the correct direction divided by the length of its migration path) is maximal for cells that start migrating at concentrations near the K d and progressively decreases at higher or lower starting concentrations. chemoattractant ͉ gradient ͉ neutrophils A n essential property of eukaryotic cells is their ability to orient in response to spatial cues. Only by correctly interpreting spatial changes in external stimuli can yeast cells mate, soil amoebae form spores, progeny of a fertilized egg form an organism, or neutrophils crawl toward their prey. Cells are known to respond to gradients of external stimuli such as chemoattractants, but how they sense and interpret gradients remains mysterious. The mystery goes beyond our ignorance of the biochemical basis of gradient sensing. More fundamentally, we have not even definitively identified the external cues sensed and interpreted by the cells and the respective roles of these cues in determining their responses to gradients.Bacteria migrate up gradients of chemoattractants, in a process called chemotaxis. Chemotaxing bacteria assess gradients temporally, by moving through the attractant concentration field, sensing the local ambient concentration, comparing the concentration at a given moment with concentrations at previous times, and changing swimming behavior accordingly (1). It has been proposed that the larger cells of eukaryotes, in contrast, sense gradients at a given moment by comparing attractant concentrations at different positions on their surfaces and thus orient themselves to crawl in the up-gradient direction by interpreting the spatial cues present in their location at that moment. In other words, such cells assess the gradient spatially and respond to purely spatial cues by directed chemotactic migration. It has been shown that both neutrophils (2) and Dictyostelium discoideum amoebae (3) can sense relatively steep gradients of chemoattractant, supplied by a micropipette, without moving and therefore without comparing ambient concentrations in different locations. In both cases, immobile cells, paralyzed by treatments that block actin polymerization, accumulate phosphatidylino...

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“…We observed the optimal chemotaxis when the mean CXCL8 concentration across the gradient was 1.25 nM. In another study where human neutrophils were assayed, the most efficient chemotaxis was induced in the microfluidic gradient device with the mean CXCL8 concentration of 3 nM across the gradient (27). Our studies with the microfluidic device show that both steepness of gradient and mean ligand concentration affect the ability of cells to persist in their migration up a gradient.…”

Section: Discussionmentioning

confidence: 48%

“…The microfluidic gradient device has several advantages over traditional methods of generating a chemokine gradient (26). With this device, stable chemokine gradients can be delivered and manipulated in a variety of ways, thereby providing a very powerful tool to study different aspects of chemotaxis, such as the effects of the chemokine concentration on chemotaxis (27). Moreover, cells can be visualized microscopically during the migration process.…”

mentioning

confidence: 99%

The IL Sequence in the LLKIL Motif in CXCR2 Is Required for Full Ligand-induced Activation of Erk, Akt, and Chemotaxis in HL60 Cells

Sai

Walker

Wikswo

et al. 2006

Journal of Biological Chemistry

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The chemotaxis of differentiated HL60 cells stably expressing CXCR2 was examined in a microfluidic gradient device where the steepness of the CXCL8 chemokine gradient was varied from 2 pg/ml/m (0 -1 ng/ml over a width of 500 m) to 50 pg/ml/m (0 -25 ng/ml over 500 m). The differentiated HL60 cells stably expressing CXCR2 exhibited little chemotaxis in response to a 0 -1 ng/ml gradient, but displayed an increasing chemotactic response as the gradient steepness increased from 0 to 5, 0 to 10, and 0 to 25 ng/ml, demonstrating that steepness of gradient is a major determinant of the relative ability of cells to persistently migrate up a chemotactic gradient. When HL60 cells expressed CXCR2 mutated in the C terminus LLKIL motif (IL to AA), ligand-induced internalization of receptors was reduced 50%, whereas cell migration along the gradient of CXCL8 was completely lost. Although both mutant and wildtype receptors could mediate Akt and Erk activation in response to CXCL8, the level of activation of these two kinases was much lower in the cell line expressing the mutant receptors. These data imply that the IL amino acid residues in the LLKIL motif are very important for activation of the signal transduction cascade, which is necessary for cells to sense the chemokine gradient and respond with chemotaxis. Moreover, because mutation of the IL residues in the LLKIL motif resulted in only 50% reduction in receptor internalization, and a 50% reduction in Akt and Erk phosphorylation, but a complete loss of chemotactic response, the data imply that IL amino acid residues in the LLKIL motif are key either for amplification or oscillation of crucial signaling events or for establishment of a threshold for signals required for chemotaxis.Chemotaxis is a process by which cells move directionally up a gradient of chemokine or chemoattractant. Chemotaxis plays an important role in the immune response through recruitment of leukocytes to the inflammatory sites (1, 2), in embryonic development where stem cells move directionally to the site of organogenesis (3), and in cancer metastasis (4 -7), whereby tumor cells preferentially chemotax to specific target organs.The phosphorylation and internalization of chemokine receptors are very important for modulation of receptor function, although the role of chemokine receptor internalization for chemotaxis remains controversial. Some studies revealed that receptor internalization was not required for certain chemoattractant receptors to mediate chemotaxis (8 -12), whereas others reported that inhibition of receptor internalization also inhibited chemotaxis (13-17). We previously reported that the CXCR2 chemokine receptor mutated in the LLKIL motif, which binds adaptor protein-2 (AP-2) 3 and heat shock 70 interacting protein (HIP), displayed compromised ligand-triggered receptor internalization and mediated impaired chemotaxis in HEK293 cells (18,19). In contrast, Richardson et al. (12) demonstrated that, in rat basophilic leukemia (RBL-2H3) cells, a C-terminal-truncated CXCR2 did not underg...

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Effective neutrophil chemotaxis is strongly influenced by mean IL-8 concentration

Cited by 127 publications

References 22 publications

Shrinky-Dink microfluidics: rapid generation of deep and rounded patterns

Shrinky-Dink microfluidics: rapid generation of deep and rounded patterns

Bound attractant at the leading vs. the trailing edge determines chemotactic prowess

The IL Sequence in the LLKIL Motif in CXCR2 Is Required for Full Ligand-induced Activation of Erk, Akt, and Chemotaxis in HL60 Cells

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