Emergence of Coordinated Collective Cell Migration under Physical Confinement

Ph.DDOCTOR OF PHILOSOPHY (NGS-MBI

Cell Culture in Microfluidic Droplets

Cell manipulation in droplets has emerged as one of the great successes of microfluidic technologies, with the development of single-cell screening. However, the droplet format has also served to go beyond single-cell studies, namely by considering the interactions between different cells or between cells and their physical or chemical environment. These studies pose specific challenges linked to the need for long-term culture of adherent cells or the diverse types of measurements associated with complex biological phenomena. Here we review the emergence of droplet microfluidic methods for culturing cells and studying their interactions. We begin by characterizing the quantitative aspects that determine the ability to encapsulate cells, transport molecules, and provide sufficient nutrients within the droplets. This is followed by an evaluation of the biological constraints such as the control of the biochemical environment and promoting the anchorage of adherent cells. This first part ends with a description of measurement methods that have been developed. The second part of the manuscript focuses on applications of these technologies for cancer studies, immunology, and stem cells while paying special attention to the biological relevance of the cellular assays and providing guidelines on improving this relevance

Cell Culture in Microfluidic Droplets

Cell manipulation in droplets has emerged as one of the great successes of microfluidic technologies, with the development of single-cell screening. However, the droplet format has also served to go beyond single-cell studies, namely by considering the interactions between different cells or between cells and their physical or chemical environment. These studies pose specific challenges linked to the need for long-term culture of adherent cells or the diverse types of measurements associated with complex biological phenomena. Here we review the emergence of droplet microfluidic methods for culturing cells and studying their interactions. We begin by characterizing the quantitative aspects that determine the ability to encapsulate cells, transport molecules, and provide sufficient nutrients within the droplets. This is followed by an evaluation of the biological constraints such as the control of the biochemical environment and promoting the anchorage of adherent cells. This first part ends with a description of measurement methods that have been developed. The second part of the manuscript focuses on applications of these technologies for cancer studies, immunology, and stem cells while paying special attention to the biological relevance of the cellular assays and providing guidelines on improving this relevance

Articulated Robot Arm for Garbage Disposal in Hospital Environment

The use of robotic arms is crucial in the medical industry, particularly in hospital settings. It can be used for a wide range of things, including as an aide in the operating room or for the removal of medical waste, among many other things. In this work, the robotic arm is designed to segregate medical waste as hazardous or non-hazardous. A dataset with five classes was created because there was no readily available data set for medical waste. The primary challenge in doing so is to programme the robotic arm’s movements and train the image dataset to classify objects as hazardous or non-hazardous. The 3D-printed robotic arm model has 6-Degree of Freedom(DOF) and is coupled with MG996R and SG90 servo motors. The robotic arm that is attached to an Arduino uno board is operated by the Blynk IoT platform. It uses YOLO V5 (You Only Look Once) algorithm to detect objects, and it favours intersection over union (IOU). To demonstrate, static robotic arm model was placed near the pile of medical waste to identify the waste and segregate it accordingly

Inference of Internal Stress in a Cell Monolayer

International audienceWe combine traction force data with Bayesian inversion to obtain an absolute estimate of the internal stress field of a cell monolayer. The method, Bayesian inversion stress microscopy, is validated using numerical simulations performed in a wide range of conditions. It is robust to changes in each ingredient of the underlying statistical model. Importantly, its accuracy does not depend on the rheology of the tissue. We apply Bayesian inversion stress microscopy to experimental traction force data measured in a narrow ring of cohesive epithelial cells, and check that the inferred stress field coincides with that obtained by direct spatial integration of the traction force data in this quasi one-dimensional geometry

High resolution microfluidic assay and probabilistic modeling reveal cooperation between T cells in tumor killing

International audienceCytotoxic T cells are important components of natural anti-tumor immunity and are harnessed in tumor immunotherapies. Immune responses to tumors and immune therapy outcomes largely vary among individuals, but very few studies examine the contribution of intrinsic behavior of the T cells to this heterogeneity. Here we show the development of a microfluidic-based in vitro method to track the outcome of antigen-specific T cell activity on many individual cancer spheroids simultaneously at high spatiotemporal resolution, which we call Multiscale Immuno-Oncology on-Chip System (MIOCS). By combining parallel measurements of T cell behaviors and tumor fates with probabilistic modeling, we establish that the first recruited T cells initiate a positive feedback loop to accelerate further recruitment to the spheroid. We also provide evidence that cooperation between T cells on the spheroid during the killing phase facilitates tumor destruction. Thus, we propose that both T cell accumulation and killing function rely on collective behaviors rather than simply reflecting the sum of individual T cell activities, and the possibility to track many replicates of immune cell-tumor interactions with the level of detail our system provides may contribute to our understanding of immune response heterogeneity

Clustering and ordering in cell assemblies with generic asymmetric aligning interactions

11 pages, 5 figuresCollective cell migration plays an essential role in various biological processes, such as development or cancer proliferation. While cell-cell interactions are clearly key determinants of collective cell migration -- in addition to individual cells self-propulsion -- the physical mechanisms that control the emergence of cell clustering and collective cell migration are still poorly understood. In particular, observations have shown that binary cell-cell collisions generally lead to anti-alignement of cell polarities and separation of pairs -- a process called contact inhibition of locomotion (CIL), which is expected to disfavor the formation of large scale cell clusters with coherent motion. Here, we adopt a joint experimental and theoretical approach to determine the large scale dynamics of cell assemblies from elementary pairwise cell-cell interaction rules. We quantify experimentally binary cell-cell interactions and show that they can be captured by a minimal equilibrium-like pairwise asymmetric aligning interaction potential that reproduces the CIL phenomenology. We identify its symmetry class, build the corresponding active hydrodynamic theory and show on general grounds that such asymmetric aligning interaction destroys large scale clustering and ordering, leading instead to a liquid-like microphase of cell clusters of finite size and short lived polarity, or to a fully dispersed isotropic phase. Finally, this shows that CIL-like asymmetric interactions in cellular systems -- or general active systems -- control cluster sizes and polarity, and can prevent large scale coarsening and long range polarity, except in the singular regime of dense confluent systems

The role of single-cell mechanical behaviour and polarity in driving collective cell migration

International audienceThe directed migration of cell collectives is essential in various physiological processes, such as epiboly, intestinal epithelial turnover, and convergent extension during morphogenesis as well as during pathological events like wound healing and cancer metastasis 1,2 . Collective cell migration leads to the emergence of coordinated movements over multiple cells. Our current understanding emphasizes that these movements are mainly driven by large-scale transmission of signals through adherens junctions 3,4 . In this study, we show that collective movements of epithelial cells can be triggered by polarity signals at the single cell level through the establishment of coordinated lamellipodial protrusions. We designed a minimalistic model system to generate one-dimensional epithelial trains confined in ring shaped patterns that recapitulate rotational movements observed in vitro in cellular monolayers and in vivo in genitalia or follicular cell rotation 5–7 . Using our system, we demonstrated that cells follow coordinated rotational movements after the establishment of directed Rac1-dependent polarity over the entire monolayer. Our experimental and numerical approaches show that the maintenance of coordinated migration requires the acquisition of a front-back polarity within each single cell but does not require the maintenance of cell-cell junctions. Taken together, these unexpected findings demonstrate that collective cell dynamics in closed environments as observed in multiple in vitro and in vivo situations 5,6,8,9 can arise from single cell behavior through a sustained memory of cell polarity