Self-assisted Amoeboid Navigation in Complex Environments

Background: Living cells of many types need to move in response to external stimuli in order to accomplish their functional tasks; these tasks range from wound healing to immune response to fertilization. While the directional motion is typically dictated by an external signal, the actual motility is also restricted by physical constraints, such as the presence of other cells and the extracellular matrix. The ability to successfully navigate in the presence of obstacles is not only essential for organisms, but might prove relevant in the study of autonomous robotic motion. Methodology/principal findings: We study a computational model of amoeboid chemotactic navigation under differing conditions, from motion in an obstacle-free environment to navigation between obstacles and finally to moving in a maze. We use the maze as a simple stand-in for a motion task with severe constraints, as might be expected in dense extracellular matrix. Whereas agents using simple chemotaxis can successfully navigate around small obstacles, the presence of large barriers can often lead to agent trapping. We further show that employing a simple memory mechanism, namely secretion of a repulsive chemical by the agent, helps the agent escape from such trapping. Conclusions/significance: Our main conclusion is that cells employing simple chemotactic strategies will often be unable to navigate through maze-like geometries, but a simple chemical marker mechanism (which we refer to as "self-assistance") significantly improves success rates. This realization provides important insights into mechanisms that might be employed by real cells migrating in complex environments as well as clues for the design of robotic navigation strategies. The results can be extended to more complicated multi-cellular systems and can be used in the study of mammalian cell migration and cancer metastasis

Concepts of GPCR-controlled navigation in the immune system

G-protein-coupled receptor (GPCR) signaling is essential for the spatiotemporal control of leukocyte dynamics during immune responses. For efficient navigation through mammalian tissues, most leukocyte types express more than one GPCR on their surface and sense a wide range of chemokines and chemoattractants, leading to basic forms of leukocyte movement (chemokinesis, haptokinesis, chemotaxis, haptotaxis, and chemorepulsion). How leukocytes integrate multiple GPCR signals and make directional decisions in lymphoid and inflamed tissues is still subject of intense research. Many of our concepts on GPCR-controlled leukocyte navigation in the presence of multiple GPCR signals derive from in vitro chemotaxis studies and lower vertebrates. In this review, we refer to these concepts and critically contemplate their relevance for the directional movement of several leukocyte subsets (neutrophils, T cells, and dendritic cells) in the complexity of mouse tissues. We discuss how leukocyte navigation can be regulated at the level of only a single GPCR (surface expression, competitive antagonism, oligomerization, homologous desensitization, and receptor internalization) or multiple GPCRs (synergy, hierarchical and non-hierarchical competition, sequential signaling, heterologous desensitization, and agonist scavenging). In particular, we will highlight recent advances in understanding GPCR-controlled leukocyte navigation by intravital microscopy of immune cells in mice

Modelling cell motility and chemotaxis with evolving surface finite elements

We present a mathematical and a computational framework for the modelling of cell motility. The cell membrane is represented by an evolving surface, with the movement of the cell determined by the interaction of various forces that act normal to the surface. We consider external forces such as those that may arise owing to inhomogeneities in the medium and a pressure that constrains the enclosed volume, as well as internal forces that arise from the reaction of the cells' surface to stretching and bending. We also consider a protrusive force associated with a reaction-diffusion system (RDS) posed on the cell membrane, with cell polarization modelled by this surface RDS. The computational method is based on an evolving surface finite-element method. The general method can account for the large deformations that arise in cell motility and allows the simulation of cell migration in three dimensions. We illustrate applications of the proposed modelling framework and numerical method by reporting on numerical simulations of a model for eukaryotic chemotaxis and a model for the persistent movement of keratocytes in two and three space dimensions. Movies of the simulated cells can be obtained from http://homepages.warwick.ac.uk/maskae/CV_Warwick/Chemotaxis.html

T Cell Migration in Three-dimensional Extracellular Matrix: Guidance by Polarity and Sensations

The locomotion of T lymphocytes within 3-D extracellular matrix (ECM) is a highly dynamic and flexible process following the principles of ameboid movement. Ameboid motility is characterized by a polarized yet simple cell shape allowing high speed, rapid directional oscillations, and low affinity interactions to the substrate that are coupled to a low degree of cytoskeletal organization lacking discrete focal contacts. At the onset of T cell migration, a default program, here described as migration-associated polarization, is initiated, resulting in the polar redistribution of cell surface receptors and cytoskeletal elements. Polarization involves protein cycling either to the leading edge (i.e. LFA-1, CD45RO, chemokine receptors, focal adhesion kinase), to a central polarizing compartment (MTOC, PKC, MARCKS), or into the uropod (CD44, CD43, ICAM- and –3, β1 integrins). The function of such compartment formation may be important in chemotactic response, scanning of encountered cells, and a flexible and adaptive interaction with the ECM itself. Due to the simple shape and a diffusely organized cytoskeleton, the interactions to the surrounding extracellular matrix are rapid and reversible and appear to allow a broad spectrum of molecular migration strategies. These range from (1) adhesive and haptokinetic following i.e. chemokine-induced motility across 2-D surfaces to (2) largely integrin-independent migration predominantly guided by shape change and morphological flexibility, as seen in 3-D type I collagen matrices. Their prominent capacity to rapidly adapt to a given structural environment coupled to contact guidance mechanisms set T cell locomotion apart from slow, focal contact-dependent and more adhesive migration strategies established by fibroblast-like cells and cell clusters. It is therefore likely that, within the tissues, besides chemotactic or haptotactic gradients, the preformed matrix structure has an important impact on T cell trafficking and positioning in health and disease

Coupled Excitable Ras and F-actin activation mediate spontaneous pseudopod formation and directed cell movement

Many eukaryotic cells regulate their mobility by external cues. Genetic studies have identified &gt;100 components that participate in chemotaxis, which hinders the identification of the conceptual framework of how cells sense and respond to shallow chemical gradients. The activation of Ras occurs during basal locomotion and is an essential connector between receptor and cytoskeleton during chemotaxis. Using a sensitive assay for activated Ras, we show here that activation of Ras and F-actin forms two excitable systems that are coupled through mutual positive feedback and memory. This coupled excitable system leads to short-lived patches of activated Ras and associated F-actin that precede the extension of protrusions. In buffer, excitability starts frequently with Ras activation in the back/side of the cell or with F-actin in the front of the cell. In a shallow gradient of chemoattractant, local Ras activation triggers full excitation of Ras and subsequently F-actin at the side of the cell facing the chemoattractant, leading to directed pseudopod extension and chemotaxis. A computational model shows that the coupled excitable Ras/F-actin system forms the driving heart for the ordered-stochastic extension of pseudopods in buffer and for efficient directional extension of pseudopods in chemotactic gradients.</p

IST Austria Thesis

Directed cell migration is a hallmark feature, present in almost all multi-cellular organisms. Despite its importance, basic questions regarding force transduction or directional sensing are still heavily investigated. Directed migration of cells guided by immobilized guidance cues - haptotaxis - occurs in key-processes, such as embryonic development and immunity (Middleton et al., 1997; Nguyen et al., 2000; Thiery, 1984; Weber et al., 2013). Immobilized guidance cues comprise adhesive ligands, such as collagen and fibronectin (Barczyk et al., 2009), or chemokines - the main guidance cues for migratory leukocytes (Middleton et al., 1997; Weber et al., 2013). While adhesive ligands serve as attachment sites guiding cell migration (Carter, 1965), chemokines instruct haptotactic migration by inducing adhesion to adhesive ligands and directional guidance (Rot and Andrian, 2004; Schumann et al., 2010). Quantitative analysis of the cellular response to immobilized guidance cues requires in vitro assays that foster cell migration, offer accurate control of the immobilized cues on a subcellular scale and in the ideal case closely reproduce in vivo conditions. The exploration of haptotactic cell migration through design and employment of such assays represents the main focus of this work. Dendritic cells (DCs) are leukocytes, which after encountering danger signals such as pathogens in peripheral organs instruct naïve T-cells and consequently the adaptive immune response in the lymph node (Mellman and Steinman, 2001). To reach the lymph node from the periphery, DCs follow haptotactic gradients of the chemokine CCL21 towards lymphatic vessels (Weber et al., 2013). Questions about how DCs interpret haptotactic CCL21 gradients have not yet been addressed. The main reason for this is the lack of an assay that offers diverse haptotactic environments, hence allowing the study of DC migration as a response to different signals of immobilized guidance cue. In this work, we developed an in vitro assay that enables us to quantitatively assess DC haptotaxis, by combining precisely controllable chemokine photo-patterning with physically confining migration conditions. With this tool at hand, we studied the influence of CCL21 gradient properties and concentration on DC haptotaxis. We found that haptotactic gradient sensing depends on the absolute CCL21 concentration in combination with the local steepness of the gradient. Our analysis suggests that the directionality of migrating DCs is governed by the signal-to-noise ratio of CCL21 binding to its receptor CCR7. Moreover, the haptotactic CCL21 gradient formed in vivo provides an optimal shape for DCs to recognize haptotactic guidance cue. By reconstitution of the CCL21 gradient in vitro we were also able to study the influence of CCR7 signal termination on DC haptotaxis. To this end, we used DCs lacking the G-protein coupled receptor kinase GRK6, which is responsible for CCL21 induced CCR7 receptor phosphorylation and desensitization (Zidar et al., 2009). We found that CCR7 desensitization by GRK6 is crucial for maintenance of haptotactic CCL21 gradient sensing in vitro and confirm those observations in vivo. In the context of the organism, immobilized haptotactic guidance cues often coincide and compete with soluble chemotactic guidance cues. During wound healing, fibroblasts are exposed and influenced by adhesive cues and soluble factors at the same time (Wu et al., 2012; Wynn, 2008). Similarly, migrating DCs are exposed to both, soluble chemokines (CCL19 and truncated CCL21) inducing chemotactic behavior as well as the immobilized CCL21. To quantitatively assess these complex coinciding immobilized and soluble guidance cues, we implemented our chemokine photo-patterning technique in a microfluidic system allowing for chemotactic gradient generation. To validate the assay, we observed DC migration in competing CCL19/CCL21 environments. Adhesiveness guided haptotaxis has been studied intensively over the last century. However, quantitative studies leading to conceptual models are largely missing, again due to the lack of a precisely controllable in vitro assay. A requirement for such an in vitro assay is that it must prevent any uncontrolled cell adhesion. This can be accomplished by stable passivation of the surface. In addition, controlled adhesion must be sustainable, quantifiable and dose dependent in order to create homogenous gradients. Therefore, we developed a novel covalent photo-patterning technique satisfying all these needs. In combination with a sustainable poly-vinyl alcohol (PVA) surface coating we were able to generate gradients of adhesive cue to direct cell migration. This approach allowed us to characterize the haptotactic migratory behavior of zebrafish keratocytes in vitro. Furthermore, defined patterns of adhesive cue allowed us to control for cell shape and growth on a subcellular scale

Modeling random crawling, membrane deformation and intracellular polarity of motile amoeboid cells

Amoeboid movement is one of the most widespread forms of cell motility that plays a key role in numerous biological contexts. While many aspects of this process are well investigated, the large cell-to-cell variability in the motile characteristics of an otherwise uniform population remains an open question that was largely ignored by previous models. In this article, we present a mathematical model of amoeboid motility that combines noisy bistable kinetics with a dynamic phase field for the cell shape. To capture cell-to-cell variability, we introduce a single parameter for tuning the balance between polarity formation and intracellular noise. We compare numerical simulations of our model to experiments with the social amoeba Dictyostelium discoideum. Despite the simple structure of our model, we found close agreement with the experimental results for the center-of-mass motion as well as for the evolution of the cell shape and the overall intracellular patterns. We thus conjecture that the building blocks of our model capture essential features of amoeboid motility and may serve as a starting point for more detailed descriptions of cell motion in chemical gradients and confined environments.Peer ReviewedPostprint (published version