Progress in Integrative Biomaterial Systems to Approach Three-Dimensional Cell Mechanotransduction.

Mechanotransduction between cells and the extracellular matrix regulates major cellular functions in physiological and pathological situations. The effect of mechanical cues on biochemical signaling triggered by cell-matrix and cell-cell interactions on model biomimetic surfaces has been extensively investigated by a combination of fabrication, biophysical, and biological methods. To simulate the in vivo physiological microenvironment in vitro, three dimensional (3D) microstructures with tailored bio-functionality have been fabricated on substrates of various materials. However, less attention has been paid to the design of 3D biomaterial systems with geometric variances, such as the possession of precise micro-features and/or bio-sensing elements for probing the mechanical responses of cells to the external microenvironment. Such precisely engineered 3D model experimental platforms pave the way for studying the mechanotransduction of multicellular aggregates under controlled geometric and mechanical parameters. Concurrently with the progress in 3D biomaterial fabrication, cell traction force microscopy (CTFM) developed in the field of cell biophysics has emerged as a highly sensitive technique for probing the mechanical stresses exerted by cells onto the opposing deformable surface. In the current work, we first review the recent advances in the fabrication of 3D micropatterned biomaterials which enable the seamless integration with experimental cell mechanics in a controlled 3D microenvironment. Then, we discuss the role of collective cell-cell interactions in the mechanotransduction of engineered tissue equivalents determined by such integrative biomaterial systems under simulated physiological conditions.

Shapes of Red Blood Cells: Comparison of 3D Confocal Images with the Bilayer-Couple Model.

Cells and organelles are shaped by the chemical and physical forces that bend cell membranes. The human red blood cell (RBC) is a model system for studying how such forces determine cell morphology. It is thought that RBCs, which are typically biconcave discoids, take the shape that minimizes their membrane-bending energies, subject to the constraints of fixed area and volume. However, recently it has been hypothesized that shear elasticity arising from the membrane-associated cytoskeleton (MS) is necessary to account for shapes of real RBCs, especially ones with highly curved features such as echinocytes. In this work we tested this hypothesis by following RBC shape changes using spherical harmonic series expansions of theoretical cell surfaces and those estimated from 3D confocal microscopy images of live cells. We found (i) quantitative agreement between shapes obtained from the theoretical model including the MS and real cells, (ii) that weakening the MS, by using urea (which denatures spectrin), leads to the theoretically predicted gradual decrease in spicule number of echinocytes, (iii) that the theory predicts that the MS is essential for stabilizing the discocyte morphology against changes in lipid composition, and that without it, the shape would default to the elliptocyte (a biconcave ellipsoid), (iv) that we were able to induce RBCs to adopt the predicted elliptocyte morphology by treating healthy discocytes with urea. The latter observation is consistent with the known connection between the blood disease hereditary elliptocytosis and spectrin mutations that weaken the cell cortex. We conclude that while the discocyte, in absence of shear, is indeed a minimum energy shape, its stabilization in healthy RBCs requires the MS, and that elliptocytosis can be explained based on purely mechanical considerations.

Coupling bending and shear effects on liposome deformation.

Cell membrane deformation induced by external mechanical stimuli has been studied extensively over the past three decades. The present study focuses on the coupling of in-plane shear H and out-of-plane bending B of liposome membrane and its influences on the deformation of a single vesicle subjected to (i) external compressive load via two parallel platens and (ii) contact forces caused by a rigid substrate. Our results show that the increase of membrane resultant stress in both loading configurations causes the liposome to become more rigid and the degree of vesicle deformation decreases when the in-plane shearing effect is dominant. A theoretical approach is developed to facilitate cell membrane characterization under different biomechanical stimuli.

Human red blood cells deformed under thermal fluid flow.

The flow-induced mechanical deformation of a human red blood cell (RBC) during thermal transition between room temperature and 42.0 degrees C is interrogated by laser tweezer experiments. Based on the experimental geometry of the deformed RBC, the surface stresses are determined with the aid of computational fluid dynamics simulation. It is found that the RBC is more deformable while heating through 37.0 degrees C to 42.0 degrees C, especially at a higher flow velocity due to a thermal-fluid effect. More importantly, the degree of RBC deformation is irreversible and becomes softer, and finally reaches a plateau (at a uniform flow velocity U > 60 microm s(-1)) after the heat treatment, which is similar to a strain-hardening dominated process. In addition, computational simulated stress is found to be dependent on the progression of thermotropic phase transition. Overall, the current study provides new insights into the highly coupled temperature and hydrodynamic effects on the biomechanical properties of human erythrocyte in a model hydrodynamic flow system.

Shape recovery of an optically trapped vesicle: effect of flow velocity and temperature.

A new biophysical approach based on optical tweezers is developed to measure the time-dependent shape transformation and recovery of a single liposome, which is induced by the sudden stop of a moving liposome from various flow velocities at constant temperature. A simple viscoelastic model has been applied to correlate the temporal geometric parameter of the deformed liposome with a characteristic time constant, i.e., the ratio of membrane viscosity to elasticity. Our results show that membrane viscosity becomes dominant in governing the shape recovery rate when sample temperature goes beyond the main phase transition temperature of the phospholipid bilayer. More importantly, flow speed and vesicle size are demonstrated as key physical determinants for the shape recovery of liposome.

Contact deformation of liposome in the presence of osmosis.

The role of osmotic pressure on the geometry of adherent liposome remains an intricate question in the mechanics of supramolecular structures. In this study, confocal reflection interference contrast microscopy in combination with cross-polarized microscopy was applied to probe the geometry of deformed liposome on fused silica substrates through the determination of a vesicle-substrate separation profile. In parallel, a theoretical model which describes the large deformation of the lipid bilayer membrane under both out-plane bending and in-plane shear forces is developed. Then, the global deformation geometry of the adherent liposome is rigorously compared with our experimental data. It is shown that the adhesion contact area increases in dimension, the liposome volume decreases, and the vesicle height decreases under the reduced osmotic pressure. The coupling of experimental data and a modified theoretical framework of the adherent liposome provides a more explicit result in comparison with previous studies and demonstrates the possibility of modeling the change of liposome mechanics under the influence of osmosis.

Thermal effect on a viscously deformed liposome in a laser trap.

The viscous drag and mechanical deformation of a single vesicle under hydrodynamics flow during the phase transition of a lipid bilayer is determined by optical tweezers experiments with the aid of computational fluid dynamics simulations. Based on the experimental geometry of the vesicle under hydrodynamics flow, the surface stresses and drag force are numerically calculated. It is found that the vesicle is less rigid and the viscous drag force of the vesicle decreases with the increase of temperature at low Reynolds number flow during sample heating. Interestingly, these mechanical properties are reversible and depend strongly on the liposome's thermotropic phase transition temperature. Overall, this study provides new insights into highly coupled thermal and hydrodynamics effects on the biomechanical properties of model membrane vesicle in physiological flow systems.