In its native form, the chemokine CX3CL1 is a firmly adhesive molecule promoting leukocyte adhesion and migration and hence involved, along with its unique receptor CX3CR1, in various inflammatory processes. Here we investigated the role of molecular aggregation in the CX3CL1 adhesiveness. Assays of bioluminescence resonance energy transfer (BRET) and homogeneous time-resolved fluorescence (HTRF) in transfected cell lines and in primary cells showed specific signals indicative of CX3CL1 clustering. Truncation experiments showed that the transmembrane domain played a central role in this aggregation. A chimera with mutations of the 12 central transmembrane domain residues had significantly reduced BRET signals and characteristics of a non-clustering molecule. This mutant was weakly adhesive according to flow and dual pipette adhesion assays and was less glycosylated than CX3CL1, although, as we demonstrated, loss of glycosylation did not affect the CX3CL1 adhesive potency. We postulate that cell surfaces express CX3CL1 as a constitutive oligomer and that this oligomerization is essential for its adhesive potency. Inhibition of CX3CL1 self-assembly could limit the recruitment of CX3CR1-positive cells and may be a new pathway for anti-inflammatory therapies.

The Surface Force Apparatus (SFA) measures directly, and with nanoscale resolution, the interaction energy vs. distance profile of planar arrays of biological molecules (e.g., lipids, polymers, or proteins). Through recent advances in the reconstitution and deposition of lipid bilayers, it is now possible to use SFA to study the interactions between membrane-incorporated biomolecules and to reveal any conformational changes and intermediate assembly states. Therein we describe two example systems. First, we show that using bilayers functionalized to carry DNA bases on their lipid headgroups, we can measure a macroscopic nucleoside–nucleoside adhesion force, from which one can obtain a molecular binding energy. Second, we describe the use of the SFA to study the interaction between SNARE proteins, which are involved in most of intracellular fusion events. Membrane fusion occurs when SNARE proteins assemble between lipid bilayers in the form of SNAREpins. SFA measurements between SNAREs embedded in lipid bilayers allowed us to elucidate the energetics and dynamics of SNAREpin folding, and to capture an intermediate binding state in SNAREpin assembly.

Polymeric capsules with a thick shell made of biodegradable and biocompatible polymer and a liquid core of perfluorooctyl bromide (PFOB) were evaluated for stability as well as for ultrasound and magnetic resonance imaging (MRI) contrast enhancement. The method of preparation allows the mean capsule diameter to be regulated between 70 nm and 25 µm and the capsule thickness-to-radius ratio from 0.25 to 0.54. Capsule diameter remains stable at 37 °C in phosphate buffer for at least 4 and 6 h for nanocapsules and microcapsules, respectively. The in vitro ultrasound signal-to-noise ratio (SNR) was measured from 40 to 60 MHz for 6 µm and 150 nm capsules: the SNR increases with capsule concentration up to 20–25 mg mL⁻¹, and then reaches a plateau that depends on capsule diameter (13.5 ± 1.5 dB for 6 µm and 6 ± 2 dB for the 150 nm capsules). The ultrasound SNR is stable for up to 20 min for microcapsules and for several hours for nanocapsules. For nanocapsules, the thinner the shell, the larger the SNR and the more compressible the capsules. Nanocapsule suspensions imaged in vitro with a commercial ultrasound imaging system (normal and tissue harmonic imaging modes, 7–14 MHz probe) were detected down to concentrations of 12.5 mg mL⁻¹. Injections of nanocapsules (200 µg ml⁻¹) in mice in vivo reveal that the initial bolus passage presents significant ultrasound enhancement of the blood pool during hepatic imaging (7–14 MHz probe, tissue harmonic imaging mode). ¹⁹F-MRI images were obtained in vitro at 9.4T using spin-echo and gradient echo sequences and allow detecting nanocapsules in suspension (50 mg mL⁻¹). In conclusion, these results show initial feasibility for development of these capsules toward a dual-modality contrast agent.

Experimentalists who measure the rupture force of a single molecular bond usually pull on that bond at a constant speed, keeping the loading rate r=dfdt constant. The challenge is to extract the energy landscape of the interaction between the two molecules involved from the experimental rupture force distribution under several loading rates. This analysis requires the use of a model for the shape of this energy landscape. Several barriers can compose the landscape, though molecular bonds with a single barrier are often observed. The Bell model is commonly used for the analysis of rupture force measurements with bonds displaying a single barrier. It provides an analytical expression of the most likely rupture force which makes it very simple to use. However, in principle, it can only be applied to landscapes with extrema whose positions do not vary under force. Here, we evaluate the general relevance of the Bell model by comparing it with another analytical model for which the landscape is harmonic in the vicinity of its extrema. Similar shapes of force distributions are obtained with both models, making it difficult to confirm the validity of the Bell model for a given set of experimental data. Nevertheless, we show that the analysis of rupture force experiments on such harmonic landscapes with the Bell model provides excellent results in most cases. However, numerical computation of the distributions of the rupture forces on piecewise-linear energy landscapes indicates that the blind use of any model such as the Bell model may be risky, since there often exist several landscapes compatible with a given set of experimental data. Finally, we derive a universal relation between the range and energy of the bond and the force spectrum. This relation does not depend on the shape of the energy landscape and can thus be used to characterize unambiguously any one-barrier landscape from experiments. All the results are illustrated with the streptavidin-biotin bond.

We describe the creation of cell adhesion mediated by cell surface engineering. The Flt3-ligand was fused to a membrane anchor made of the diphtheria toxin translocation domain. The fusion protein was attached to the surface of a cell by an acid pulse. Contact with another cell expressing the receptor Flt3 lead to its activation. This activity involved direct cell–cell contact. A mean force of 20 nN was needed to separate functionalized cells after 5 min of contact. Overall, we showed that it is possible to promote specific cell–cell adhesion by attaching protein ligands at the surface of cells.