Physiological ramifications of constrained collective cell migration.

Constraining collective cell migration in vitro using different types of engineered substrates such as microstructured surfaces or adhesive patterns of different shapes and sizes often leads to the emergence of specific patterns of motion. Recently, analogies between the behavior of cellular assemblies and that of active fluids have enabled significant advances in our understanding of collective cell migration; however, the physiological relevance and potential functional consequences of the resulting migration patterns remain elusive. Here we describe the different patterns of collective cell migration that have been reported in vitro in response to geometrical constraints, explore the in vivo pertinence of the in vitro systems used to impose the geometrical constraints, and discuss the potential physiological ramifications of the collective migration patterns that emerge as a result of physical constraints. We conclude by highlighting key upcoming challenges in the exciting field of constrained collective cell migration.

Topography-induced large-scale antiparallel collective migration in vascular endothelium.

Collective migration of vascular endothelial cells is central for embryonic development, angiogenesis, and wound closure. Although physical confinement of cell assemblies has been shown to elicit specific patterns of collective movement in various cell types, endothelial migration in vivo often occurs without confinement. Here we show that unconfined endothelial cell monolayers on microgroove substrates that mimic the anisotropic organization of the extracellular matrix exhibit a specific type of collective movement that takes the form of a periodic pattern of antiparallel cell streams. We further establish that the development of these streams requires intact cell-cell junctions and that stream sizes are particularly sensitive to groove depth. Finally, we show that modeling the endothelial cell sheet as an active fluid with the microgrooves acting as constraints on cell orientation predicts the occurrence of the periodic antiparallel cell streams as well as their lengths and widths. We posit that in unconfined cell assemblies, physical factors that constrain or bias cellular orientation such as anisotropic extracellular matrix cues or directed flow-derived shear forces dictate the pattern of collective cell movement.

Integration of substrate- and flow-derived stresses in endothelial cell mechanobiology.

Endothelial cells (ECs) lining all blood vessels are subjected to large mechanical stresses that regulate their structure and function in health and disease. Here, we review EC responses to substrate-derived biophysical cues, namely topography, curvature, and stiffness, as well as to flow-derived stresses, notably shear stress, pressure, and tensile stresses. Because these mechanical cues in vivo are coupled and are exerted simultaneously on ECs, we also review the effects of multiple cues and describe burgeoning in vitro approaches for elucidating how ECs integrate and interpret various mechanical stimuli. We conclude by highlighting key open questions and upcoming challenges in the field of EC mechanobiology.

Is there a universal mechanism of cell alignment in response to substrate topography?

Cell alignment and elongation in the direction of anisotropic and aligned topographies are key manifestations of cellular contact guidance and are observed in many cell types. Whether this observation occurs through a universal mechanism remains to be established. In this Views article, we begin by presenting the most widely accepted model of topography-driven cell alignment which posits that anisotropic topographies impose lateral constraints on the growth of focal adhesions and actin stress fibers, thereby driving anisotropic force generation and cellular elongation and alignment. We then discuss particular scenarios where alternative or complementary mechanisms of cell alignment appear to be at play. These include the cases of specific cell types such as amoeboid-like cells and neurons as well as certain topography sizes. Finally, we review the role of the actin cytoskeleton in modulating topography-driven cell alignment and underscore the need for elucidating the role that other cytoskeletal elements play. We close by identifying key open questions the responses to which will significantly enhance our understanding of the role of cellular contact guidance in health and disease.

Cellular and Subcellular Contact Guidance on Microfabricated Substrates.

Topography of the extracellular environment is now recognized as a major biophysical regulator of cell behavior and function. The study of the influence of patterned substrates on cells, named contact guidance, has greatly benefited from the development of micro and nano-fabrication techniques, allowing the emergence of increasingly diverse and elaborate engineered platforms. The purpose of this review is to provide a comprehensive view of the process of contact guidance from cellular to subcellular scales. We first classify and illustrate the large diversity of topographies reported in the literature by focusing on generic cellular responses to diverse topographical cues. Subsequently, and in a complementary fashion, we adopt the opposite approach and highlight cell type-specific responses to classically used topographies (arrays of pillars or grooves). Finally, we discuss recent advances on the key subcellular and molecular players involved in topographical sensing. Throughout the review, we focus particularly on neuronal cells, whose unique morphology and behavior have inspired a large body of studies in the field of topographical sensing and revealed fascinating cellular mechanisms. We conclude by using the current understanding of the cell-topography interactions at different scales as a springboard for identifying future challenges in the field of contact guidance.

The basement membrane as a structured surface - role in vascular health and disease.

The basement membrane (BM) is a thin specialized extracellular matrix that functions as a cellular anchorage site, a physical barrier and a signaling hub. While the literature on the biochemical composition and biological activity of the BM is extensive, the central importance of the physical properties of the BM, most notably its mechanical stiffness and topographical features, in regulating cellular function has only recently been recognized. In this Review, we focus on the biophysical attributes of the BM and their influence on cellular behavior. After a brief overview of the biochemical composition, assembly and function of the BM, we describe the mechanical properties and topographical structure of various BMs. We then focus specifically on the vascular BM as a nano- and micro-scale structured surface and review how its architecture can modulate endothelial cell structure and function. Finally, we discuss the pathological ramifications of the biophysical properties of the vascular BM and highlight the potential of mimicking BM topography to improve the design of implantable endovascular devices and advance the burgeoning field of vascular tissue engineering.

Topographical cues control the morphology and dynamics of migrating cortical interneurons.

In mammalian embryos, cortical interneurons travel long distances among complex three-dimensional tissues before integrating into cortical circuits. Several molecular guiding cues involved in this migration process have been identified, but the influence of physical parameters remains poorly understood. In the present study, we have investigated in vitro the influence of the topography of the microenvironment on the migration of primary cortical interneurons released from mouse embryonic explants. We found that arrays of PDMS micro-pillars of 10 µm size and spacing, either round or square, influenced both the morphology and the migratory behavior of interneurons. Strikingly, most interneurons exhibited a single and long leading process oriented along the diagonals of the square pillared array, whereas leading processes of interneurons migrating in-between round pillars were shorter, often branched and oriented in all available directions. Accordingly, dynamic studies revealed that growth cone divisions were twice more frequent in round than in square pillars. Both soma and leading process tips presented forward directed movements within square pillars, contrasting with the erratic trajectories and more dynamic movements observed among round pillars. In support of these observations, long interneurons migrating in square pillars displayed tight bundles of stable microtubules aligned in the direction of migration. Overall, our results show that micron-sized topography provides global spatial constraints promoting the establishment of different morphological and migratory states. Remarkably, these different states belong to the natural range of migratory behaviors of cortical interneurons, highlighting the potential importance of topographical cues in the guidance of these embryonic neurons, and more generally in brain development.

In Vitro Models to Analyze the Migration of MGE-Derived Interneurons.

In the developing brain, MGE-derived interneuron precursors migrate tangentially long distances to reach the cortex in which they later establish connections with the principal cortical cells to control the activity of adult cortical circuits. Interneuron precursors exhibit complex morphologies and migratory properties, which are difficult to study in the heterogeneous and uncontrolled in vivo environment. Here, we describe two in vitro models in which the migration environment of interneuron precursors is significantly simplified and where their migration can be observed for one to 3 days. In one model, MGE-derived interneuron precursors are cultured and migrate on a flat synthetic substrate. In the other model, fluorescent MGE-derived interneuron precursors migrate on a monolayer of dissociated cortical cells. In both models, cell movements can be recorded by time-lapse microscopy for dynamic analyses.

Cortical interneurons migrating on a pure substrate of N-cadherin exhibit fast synchronous centrosomal and nuclear movements and reduced ciliogenesis.

The embryonic development of the cortex involves a phase of long distance migration of interneurons born in the basal telencephalon. Interneurons first migrate tangentially and then reorient their trajectories radially to enter the developing cortex. We have shown that migrating interneurons can assemble a primary cilium, which maintains the centrosome to the plasma membrane and processes signals to control interneuron trajectory (Baudoin et al., 2012). In the developing cortex, N-cadherin is expressed by migrating interneurons and by cells in their migratory pathway. N-cadherin promotes the motility and maintains the polarity of tangentially migrating interneurons (Luccardini et al., 2013). Because N-cadherin is an important factor that regulates the migration of medial ganglionic eminence (MGE) cells in vivo, we further characterized the motility and polarity of MGE cells on a substrate that only comprises this protein. MGE cells migrating on a N-cadherin substrate were seven times faster than on a laminin substrate and two times faster than on a substrate of cortical cells. A primary cilium was much less frequently observed on MGE cells migrating on N-cadherin than on laminin. Nevertheless, the mature centriole (MC) frequently docked to the plasma membrane in MGE cells migrating on N-cadherin, suggesting that plasma membrane docking is a basic feature of the centrosome in migrating MGE cells. On the N-cadherin substrate, centrosomal and nuclear movements were remarkably synchronous and the centrosome remained near the nucleus. Interestingly, MGE cells with cadherin invalidation presented centrosomal movements no longer coordinated with nuclear movements. In summary, MGE cells migrating on a pure substrate of N-cadherin show fast, coordinated nuclear and centrosomal movements, and rarely present a primary cilium.