Abstract
Cellular locomotion is a central hallmark of eukaryotic life. It is governed by cell-extrinsic molecular factors, which can either emerge in the soluble phase or as immobilized, often adhesive ligands. To encode for direction, every cue must be present as a spatial or temporal gradient. Here, we developed a microfluidic chamber that allows measurement of cell migration in combined response to surface immobilized and soluble molecular gradients. As a proof of principle we study the response of dendritic cells to their major guidance cues, chemokines. The majority of data on chemokine gradient sensing is based on in vitro studies employing soluble gradients. Despite evidence suggesting that in vivo chemokines are often immobilized to sugar residues, limited information is available how cells respond to immobilized chemokines. We tracked migration of dendritic cells towards immobilized gradients of the chemokine CCL21 and varying superimposed soluble gradients of CCL19. Differential migratory patterns illustrate the potential of our setup to quantitatively study the competitive response to both types of gradients. Beyond chemokines our approach is broadly applicable to alternative systems of chemo- and haptotaxis such as cells migrating along gradients of adhesion receptor ligands vs. any soluble cue.
Highlights
IntroductionOnce in the lymph node, the cells experience a second chemokine, (C-C motif) ligand[19] (CCL19), which interacts with the same receptor (C-C chemokine receptor 7, CCR7) but interacts only weakly with sugars
Lymphatic vessels, from where they are flushed into the sinus of lymph nodes
To quantitatively track immune cell migration in simultaneous response to chemotactic and haptotactic gradients we developed a microfluidic device that allows (i) patterning of bound chemokine gradients, (ii) precise positioning of immune cells on these haptotactic gradients and (iii) the generation of diffusion-based soluble chemokine gradients superimposed on haptotactic gradients in small microfluidic migration chambers
Summary
Once in the lymph node, the cells experience a second chemokine, (C-C motif) ligand[19] (CCL19), which interacts with the same receptor (C-C chemokine receptor 7, CCR7) but interacts only weakly with sugars. It has been shown in vitro that the directionality of DCs migrating on homogenously immobilized CCL21 can be biased by gradients of soluble CCL1910. How DCs respond to immobilized and co-existing immobilized and soluble chemokine gradients remains elusive. We developed an in vitro setup to study the significance and interaction of co-existing bound and soluble chemokine gradients for directed cell migration. To this end we engineered a microfluidic device to generate diffusion-based chemokine gradients, which allows simultaneous surface-immobilization of arbitrarily graded chemokine patterns. We used DCs as a model to track migration in response to soluble and immobilized chemokine on a single cell level in real time
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