Shaping cells to control their activation and mechanics

We introduced the use of “shaped T cells“ during the thesis of A. Sadoun, since the spreading state of cells, on any adhesive molecule, is strongly inhomogeneous, giving rise to strong dispersion of the mechanical parameters at population levels . The cells were adhered on micro-stamped adhesive / repulsive patterns that will impose given levels of adhesion and cell shapes. The design of the patterning strategy was crucial since  the T cells are small (~10 µm), and extremely activable, so required specific passivation strategies : using an advanced calcium imaging processing technique at cell scale, we  determined the best adhesive / repulsive molecular combination, the most favourable concentration conditions and cell densities, in order to achieve different patterned, non activated but activable T cells. We also obtained patterned model APCs (transfected COS-7 cells, see below). All cells were characterized mechanically using AFM indentation. We and others have established that APC like cells (monocytes, macrophages, COS-7) are one order of magnitude stiffer (~1kPa) than typical T cells (~100Pa).  We quantified, using RICM and confocal microscopies the adhesion and structures of the patterned cells (membrane, cytoskeleton, nucleus) and their activation by measureing their calcium fluxes when submitted to soluble antiCD3 or sedimenting model APCs. As such, we established a platform of standardized T cells for mechano-activation studies. We also studied the structure and interaction of immune cells with standardized APC cells, which we caracterized using a collection of optical microscopies, surface microscopies and AFM imaging and force modes.

Structures are also developped in the frame of the FR/MEX IRP BioPhysImmuno (Coord. PH Puech)

Fig: Patterned T cells as imaged in confocal (left) and mechanically tested using AFM colloidal force probe (right). The image is the superposition of fluorescence (for the patterns, 15µm in diameter) and transmission microscopies.

Controlling T cells shape, mechanics and activation by micropatterning. A. Sadoun, M. Biarnes-Pelicot, L. Ghesquiere-Dierickx, A. Wu, O. Théodoly, L. Limozin, Y. Hamon, P.-H. Puech, Sci Rep. 2021 Mar 24;11(1):6783. doi: 10.1038/s41598-021-86133-1