Yet another summer that ends with the graduation of a new PhD. Nicolas Garcia Seyda joined the lab as an engineer before deciding to pursue his PhD in the sunny city. He successfully defended his thesis entitled “Live imaging of leukocyte migration in controlled in vitro setups”. His presence in the lab is surely going to be missed, we wish him the best on his new adventures!
The interactions between haematopoietic and stromal cells are profoundly altered by leukaemias, contributing to the phenomena of resistance to myeloablative treatments. In this study, we followed the dynamics of JAM adhesion molecules at the membrane between leukaemic and stromal cells by videonanoscopy in order to study the establishment and evolution of these cellular junctions. The trajectories of JAMs were analysed with near-nanometer precision using a dedicated MTT (Multi-Target Tracing, Sergé et al. Nature Methods 2008) algorithm extended to 2 colours, which allows to reveal the signature of interaction and stabilization events at cell contacts. We have thus characterised the involvement of JAMs in the interaction mechanisms of tumour cells as well as the maintenance of stem cells in bone marrow niches through enhanced interaction. From a therapeutic perspective, we destabilised leukaemic stem cells using blocking antibodies opening opportunities for disrupting LSC resistance mechanisms.
https://doi.org/10.1242/jcs.258736
(B) Maximum projection of a 500-frame videonanoscopy acquisition, showing JAM-B and JAM-C positions over time (left). Maps of JAM-B and JAM-C trajectories represented by gradients of green and magenta, respectively, according to time, and superimposed on the transmission image of the cells (right). Inserts show magnifications of the framed areas. Spatiotemporal colocalizations are denoted by white circles with a size proportional to duration. Several concentric circles correspond to successive colocalizations at a nearby locations but with different durations. (C) Images from the same videonanoscopy acquisition corresponding to the area framed in B, with colocalization events or not (white circle or green/magenta arrowheads, respectively).
Dr. Cecile Sykes “Active cell deformation through actinassembly, and how we may infer nucleus deformation” Amphi 12
This article presents the evidence that immune cells are regulating very rapidely, even before classical signalling times as recorded by calcium fluxes, their mechanical properties when encountering an activating substrate, being either artificial (bead) or physiological (APC). For this, we developed a micropipette-based rheometer to track cell viscous and elastic properties. We have shown that leukocytes become up to 10 times stiffer and more viscous during their activation. Elastic and viscous properties evolve in parallel, preserving a ratio characteristic of the leukocyte subtype. These mechanical measurements set up a complete picture of the mechanics of leukocyte activation and provide a signature of cell function
Activation of three types of leukocytes studied with the micropipette rheometer. Left (a): T cell, middle (b): B cell, right (c): PLB cells. All bars represent 5 μm.
Geoffrey Delhaye has successfully defended his PhD thesis! After 3 years in sunny Marseille, he’s off to a new adventure befitting his free spirit. Our warmest congratulations to you, Geoffrey, and best of luck in your future pursuits!
Prof. Primoz Ziherl “Surface-tension based models of epithelia” (coorganized with Centuri) Hexagone
LAI is happy to welcome the new batch of PhDs joining us this year.
Valentine Seveau will be using micro-fluidic techniques and micro-patterning technologies (PRIMO Alveole) to study the guidance of T cell lymphocytes mediated by integrins and cellular adhesion molecules on a substrate, also known as haptotaxis. She’ll be co-directed by Olivier Théodoly and Marie-Pierre Valignat.
Elodie Lafargue will be trying to characterize for the first time the rest structure of voltage-dependent channels at the molecular level as well as the dynamics of electrical signal transduction when opening or closing the transmembrane channel thanks to two novel breakthroughs in High-Speed Atomic Force Microscopy (HS-AFM) technology. Elodie will be supervised by Ignacio Casuso.
Rémy Torro and Lama Awada also joined us through the CENTURI PhD program. Rémy will be co-directed by Laurent Limozin at LAI and Kheya Sengupta at CINAM, he’ll be using reflection interference contrast microscopy (RICM) and deep learning-based techniques, as a systematic approach to perform 3D reconstructions of cell membranes at the interface with substrates. Lama on the other hand, is supervised by Philippe Robert at LAI and Pierre Milpied at CIML and she’ll be trying to characterize the nature of physical cues driving antibody affinity maturation in B cell immune responses using the laminar flow chamber in 2D while simultaneously integrating parallel analysis of transcriptome and antibody genes sequences at single-cell resolution.
Welcome everyone and we wish you a successful journey!
SIGNIFICANCE Leukocytes have a ubiquitous capacity to migrate on or in solid matrices and with or without adhesion, which is instrumental to fight infections. The precise mechanisms sustaining migration remain, however, arguable. It is for instance widely accepted that leukocytes cannot crawl on two-dimensional substrates without adhesion. In contrast, we showed that human lymphocytes swim on nonadherent two-dimensional substrates and in suspension. Furthermore, our experiments and modeling suggest that propulsion hardly rely on cell body deformations and predominantly on molecular paddling by transmembrane proteins protruding outside the cell. For physics, this study reveals a new type of microswimmer, and for biology, it suggests that leukocyte’s ubiquitous crawling may have evolved from an early machinery of swimming shared by various eukaryotic cells.
Biophysical Journal 119, 1157–1177, September 15, 2020 1157
This paper was commented on in Science (link), CellPress (link), Eurekalert (link) and Science&Vie (link) and CNRS (link).
Cell guidance by anchored molecules, or haptotaxis, is crucial in development, immunology and cancer. Adhesive haptotaxis, or guidance by adhesion molecules, is well established for mesenchymal cells such as fibroblasts, whereas its existence remains unreported for amoeboid cells that require less or no adhesion in order to migrate. We show that, in vitro, amoeboid human T lymphocytes develop adhesive haptotaxis mediated by densities of integrin ligands expressed by high endothelial venules. Moreover, lymphocytes orient towards increasing adhesion with VLA4 integrins (also known as integrin α4β1), like all mesenchymal cells, but towards decreasing adhesion with LFA-1 integrins (also known as integrin αLβ4), which has not previously been observed. This counterintuitive ‘reverse haptotaxis’ cannot be explained by existing mechanisms of mesenchymal haptotaxis involving either competitive anchoring of cell edges under tension or differential integrin-activated growth of lamellipodia, because they both favor orientation towards increasing adhesion. The mechanisms and functions of amoeboid adhesive haptotaxis remain unclear; however, multidirectional integrin-mediated haptotaxis might operate around transmigration ports on endothelia, stromal cells in lymph nodes, and inflamed tissue where integrin ligands are spatially modulated.
https://doi.org/10.1242/jcs.24288
Left : T cells on repetition of adhesion gradients of integrin lagands. Right- T cells migrate toward lower adhesion on ICAM-1 lignads.
Dalia El Arawi started her LAI journey during COVID19 lockdown on the physical and computational aided design of bispecific antibodies for cancer immunotherapy under the supervision of Laurent Limozin. Her main objective will consist of testing bispecific antibodies using in vitro Antibody Dependent Cell Cytotoxicity mediated by NK cells employing TIRFM and RICM.
Adhesion & Inflammation Lab 
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