A landmark in vivo study of tissue navigation (Weber et al., Science 2013) showed that mouse skin contains endogenous, steeply decaying gradients of the chemokine CCL21 immobilized on extracellular-matrix heparan sulfate around lymphatic vessels, and that these haptotactic gradients — rather than soluble chemokine gradients — direct migrating dendritic cells into afferent lymphatics. The study left open whether this is a general principle of interstitial cell guidance: does haptotaxis along immobilized chemokine gradients operate for other chemokine–receptor systems, in other tissues, and for endogenous leukocyte subsets during inflammation; how are such gradients quantitatively established and maintained in intact tissue (production rates, diffusion, matrix binding, proteolytic cleavage, scavenging); what is the role of the second CCR7 ligand CCL19; and how much does haptotaxis contribute functionally relative to soluble gradients, chemokinesis, and mechanical cues? A decade of follow-up has substantially advanced the CCL21/CCR7 system itself — quantifying gradient-sensing thresholds, revealing unexpected independence of gradient shape from lymphatic heparan sulfates, identifying ACKR4-mediated scavenging and uPA/plasmin-mediated cleavage as gradient-remodeling mechanisms, and extending CCL21 guidance to intralymphatic crawling and transmigration — but no in vivo demonstration of chemokine haptotaxis for another chemokine–receptor system or tissue has been reported, and a quantitative biophysical account of gradient formation in intact tissue remains incomplete. The problem remains open.

Background

Dendritic cells (DCs) are antigen-presenting cells that patrol peripheral tissues, take up antigen, and carry it through afferent lymphatic vessels (the thin-walled vessels that drain tissue fluid toward lymph nodes) to initiate adaptive immune responses. To make this journey, a DC must first migrate directionally through the interstitium — the collagen- and glycosaminoglycan-rich extracellular space between cells — to find a lymphatic capillary. Chemokines are small secreted proteins that control leukocyte positioning by activating G-protein-coupled receptors. The receptor CCR7 and its two ligands, CCL19 and CCL21, govern DC trafficking to lymph nodes. A key biochemical difference between the two ligands is that CCL21 carries a positively charged C-terminal tail that binds glycosaminoglycans (GAGs), the sugar chains such as heparan sulfate that decorate cell surfaces and the extracellular matrix, and this binding immobilizes CCL21 on tissue structures. Classical chemotaxis is directed migration up a gradient of a soluble cue; chemokinesis is undirected stimulation of motility by a uniformly distributed cue; haptotaxis is directed migration along a gradient of a substrate-bound (immobilized) cue, a concept introduced originally for gradients of adhesion molecules. Before 2013, in vivo chemokine gradients in tissue parenchyma had rarely been visualized directly, and the best-established in vivo chemokine function — triggering leukocyte arrest on vessel walls during exit from the bloodstream — does not involve gradients at all. In vitro, Schumann et al. (Immunity 2010) had shown that DCs respond to immobilized CCL21 fields in microfabricated chambers. Weber et al. (Science 2013), using split mouse-ear explants and exogenously added mature DCs, then provided the first functional in vivo demonstration: quantitative imaging revealed CCL21 depots inside lymphatic endothelial cells and steeply decaying gradients of CCL21 immobilized on heparan sulfates in the perilymphatic interstitium; DCs approached lymphatic vessels directionally from up to about 90 micrometers away; and experimentally delocalizing the immobilized CCL21, or swamping the endogenous gradient with excess soluble ligand, abolished directed migration. CCL19 was dispensable for vessel entry in this system. The authors concluded that haptotaxis along immobilized chemokine gradients is functionally established in tissues and suggested, because many chemokines bind GAGs, that it could be a widely used guidance principle. That demonstration, however, came from one chemokine–receptor system, one tissue, and exogenously supplied cells, and the biophysical processes building the gradient were not quantitatively characterized. Subsequent work has refined this picture in the CCL21/CCR7 system itself: Schwarz et al. (2017) quantified in vitro how DCs read immobilized CCL21 gradients, finding directionality governed by the signal-to-noise ratio of CCL21–CCR7 binding (a product of absolute concentration and gradient steepness) with the in vivo measured gradients matching optimal sensing conditions; Vaahtomeri et al. (2021) found, surprisingly, that interstitial CCL21 gradient shape and function are largely independent of heparan sulfates produced by lymphatic endothelium, leaving the gradient-formation mechanism unresolved; the atypical chemokine receptor ACKR4 (a non-signaling scavenger receptor, i.e., a receptor that internalizes its ligand and degrades it without provoking migratory signaling) shapes CCL21/CCL19 distributions in lymph nodes and skin; and Collado-Diaz et al. (2026) showed that urokinase-plasminogen-activator-dependent plasmin cleavage of CCL21 generates a soluble truncated variant (CCL21-ΔC) with enhanced gradient-forming capacity, a process enhanced during inflammation that regulates DC entry into lymphatics. In parallel, later steps of the same journey were shown to be CCL21-guided: intralymphatic DC crawling (Russo et al. 2016) and transmigration at lymphatic endothelial multicellular junctions via spatially targeted CCL21 exocytosis (Liaqat et al. 2024). Reviews of chemokine presentation by glycosaminoglycans (Graham, Handel and Proudfoot 2019) argue that matrix-bound chemokine display is broadly important, yet direct in vivo demonstrations of haptotactic guidance remain confined to the CCL21/CCR7 system in skin and lymphoid tissues, and in type 2 inflammation DCs migrate to draining lymph nodes even when CCR7 is low, supported by bioactive lipid mediators (Meloun and León 2025), showing that parallel guidance pathways can operate.

Problem Statement

The overarching objective is to establish whether haptotaxis — directed cell migration along gradients of a chemokine (a small secreted protein that controls leukocyte positioning by activating G-protein-coupled receptors) immobilized on extracellular matrix and cell-surface glycosaminoglycans (GAGs, the sugar chains such as heparan sulfate that decorate cell surfaces and the extracellular matrix) — is a general principle of in vivo tissue navigation, and how such gradients are quantitatively built and maintained. Building on the demonstration by Weber et al. 2013 that endogenous, heparan-sulfate-immobilized gradients of the chemokine CCL21 in mouse skin guide dendritic cells to lymphatic vessels, the following linked questions remain: (1) Generality across systems and tissues: does the same haptotactic mechanism direct cells in vivo for chemokine–receptor systems other than CCL21/CCR7 (CCL21 and CCL19 are the two ligands of the receptor CCR7, which governs dendritic-cell trafficking to lymph nodes), and in tissues other than skin and lymphoid organs? This requires visualizing endogenous immobilized chemokine gradients in the target tissue and showing that their experimental disruption abolishes directed migration of the resident cell type. (2) Generality across physiological states and cell types: does the mechanism operate for endogenous dendritic cells (not exogenously transferred cells) during inflammation, where gradients are actively remodeled, and for other endogenous leukocyte subsets? (3) Biophysical mechanism: a quantitative account of how immobilized chemokine gradients are established and maintained in intact tissue — production and release rates from the producing cells, diffusion in the interstitium, glycosaminoglycan binding and unbinding kinetics, proteolytic cleavage, and scavenging by atypical chemokine receptors (non-signaling scavenger receptors that internalize and degrade their chemokine ligand without provoking migratory signaling) — sufficient to predict gradient shape and its response to perturbation. (4) Context-dependent interplay: the role of the soluble CCR7 ligand CCL19 in settings where it matters, and the functional weighting of haptotaxis relative to soluble-gradient chemotaxis (directed migration up a gradient of a dissolved cue), chemokinesis (undirected stimulation of motility by a uniformly distributed cue), mechanical confinement and flow, and lipid-mediator guidance in complex tissue environments. Answers may establish the mechanism for specific additional chemokine–receptor systems or tissues, or demonstrate its absence there; a complete answer characterizes the scope, mechanism, and relative importance of haptotactic guidance in interstitial cell navigation.

References

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