Neutrophil Migration in Opposing Chemoattractant Gradients Using Microfluidic Chemotaxis Devices

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

doi:10.1007/s10439-005-2503-6

Cited by 87 publications

(91 citation statements)

References 23 publications

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“…Chemokine gradients can be sensed by cells in a number of ways (36,37), and CCR7-driven DC chemotaxis in 3D has not to date been quantitatively described. We sought to quantitate and compare DC chemotactic behavior in 3D in different gradients of CCL21 and CCL19, considering that the absolute concentration gradient (∇C), average concentration (C Avg ), and percent gradient (∇C∕C Avg ) may all contribute to how DCs sense and respond to the chemokine.…”

Section: Resultsmentioning

confidence: 99%

Dendritic cell chemotaxis in 3D under defined chemokine gradients reveals differential response to ligands CCL21 and CCL19

Haessler

Pisano

et al. 2011

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

181

184

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Dendritic cell (DC) homing to the lymphatics and positioning within the lymph node is important for adaptive immunity, and is regulated by gradients of CCL19 and CCL21, ligands for CCR7. Despite the importance of DC chemotaxis, it is not well understood how DCs interpret gradients of these chemokines in a complex 3D microenvironment. Here, we use a microfluidic device that allows rapid establishment of stable gradients in 3D matrices to show that DC chemotaxis in 3D can respond to CCR7 ligand gradients as small as 0.4%, which helps explain how DCs sense lymphatic vessels in an environment where broadcast distance for chemokine diffusion is hindered by convective flows into the vessel. Interestingly, DCs displayed similar sensitivities to both chemokines at small gradients (≤60 nM∕mm), but migrated more efficiently towards higher gradients of CCL21, which unlike CCL19 binds strongly to matrix proteoglycans and signals without the need for internalization. Furthermore, cells preferentially migrated towards CCL21 when exposed to equal and opposite gradients of CCL21 and CCL19 simultaneously, even when matrix-binding of CCL21 was prevented. Although these ligands have similar binding affinity to CCR7, our results demonstrate that, in a 3D environment, CCL21 is a more potent directional cue for DC migration than CCL19. These findings provide new quantitative insight into DC chemotaxis in a physiological 3D environment and suggest how CCL19 and CCL21 may signal differently to fine-tune DC homing and positioning within the lymphatic system. These results also have broad relevance to other systems of cell chemotaxis, which remain poorly understood in the 3D context.D endritic cells (DCs) are considered the most potent and professional antigen-presenting cells. DCs are positioned throughout the periphery, and when activated, migrate to lymphatic vessels and into lymph nodes, where they can direct antigen-specific Tcell responses. Upon activation or maturation, DCs upregulate the C-C chemokine-receptor CCR7, which allows them to sense and home towards CCR7 ligand-secreting lymphatic vessels and lymph nodes (1). The two known ligands for CCR7 are the C-C chemokines CCL21 and CCL19 (2); both are secreted by stromal cells in the lymph node paracortex to properly position DCs with CCR7 þ naïve T cells for their activation. CCL21, but not CCL19, is also expressed by the endothelium of lymphatic vessels in the periphery (1). Thus, CCR7-mediated chemotaxis is critical for DC homing to, and positioning within, lymph nodes and for T cell activation there (3).CCL19 and CCL21 function as directional signals presumably by virtue of concentration gradients (∇C) that guide DCs towards areas of increasing concentrations. Interestingly, CCL19 and CCL21 have similar binding affinities for CCR7 (4-6) and similar chemotactic potential for DCs and T cells under 2D conditions (7,8), but differ in their internalization (8, 9) as well as their binding affinity to extracellular matrix (ECM) proteoglycans. Specifically, the positively charged C t...

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Section: Resultsmentioning

confidence: 99%

Dendritic cell chemotaxis in 3D under defined chemokine gradients reveals differential response to ligands CCL21 and CCL19

Haessler

Pisano

et al. 2011

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

181

184

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“…Although neutrophils have been studied extensively using microfluidic chambers (29,30), Lin and Butcher (31) were the first to extend the use of simple microfluidic gradients to T cells. In their study, T cells were presented with overlapping microfluidic gradients of two homeostatic chemokines: CCL19 and CXCL12.…”

mentioning

confidence: 99%

Dendritic Cells Distinguish Individual Chemokine Signals through CCR7 and CXCR4

Ricart¹,

John

Lee

et al. 2011

The Journal of Immunology

129

143

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Dendritic cells (DCs) respond to chemotactic signals to migrate from sites of infection to secondary lymphoid organs where they initiate the adaptive immune response. The key chemokines directing their migration are CCL19, CCL21, and CXCL12, but how signals from these chemokines are integrated by migrating cells is poorly understood. Using a microfluidic device, we presented single and competing chemokine gradients to murine bone-marrow derived DCs in a controlled, time-invariant microenvironment. Experiments performed with counter-gradients revealed that CCL19 is 10–100-fold more potent than CCL21 or CXCL12. Interestingly, when the chemoattractive potencies of opposing gradients are matched, cells home to a central region in which the signals from multiple chemokines are balanced; in this region, cells are motile but display no net displacement. Actin and myosin inhibitors affected the speed of crawling but not directed motion, whereas pertussis toxin inhibited directed motion but not speed. These results provide fundamental insight into the processes that DCs use to migrate toward and position themselves within secondary lymphoid organs.

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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.…”

mentioning

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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Neutrophil Migration in Opposing Chemoattractant Gradients Using Microfluidic Chemotaxis Devices

Cited by 87 publications

References 23 publications

Dendritic cell chemotaxis in 3D under defined chemokine gradients reveals differential response to ligands CCL21 and CCL19

Dendritic cell chemotaxis in 3D under defined chemokine gradients reveals differential response to ligands CCL21 and CCL19

Dendritic Cells Distinguish Individual Chemokine Signals through CCR7 and CXCR4

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

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