Various signals in tissue microenvironments are often unevenly distributed around cells. Cellular responses to asymmetric cell‐matrix adhesion in a 3D space remain generally unclear and are to be studied at the single‐cell resolution. Here, the authors developed a droplet‐based microfluidic approach to manufacture a pure population of single cells in a microscale layer of compartmentalized 3D hydrogel matrices with a tunable spatial presentation of ligands at the subcellular level. Cells elongate with an asymmetric presentation of the integrin adhesion ligand Arg‐Gly‐Asp (RGD), while cells expand isotropically with a symmetric presentation of RGD. Membrane tension is higher on the side of single cells interacting with RGD than on the side without RGD. Finite element analysis shows that a non‐uniform isotropic cell volume expansion model is sufficient to recapitulate the experimental results. At a longer timescale, asymmetric ligand presentation commits mesenchymal stem cells to the osteogenic lineage. Cdc42 is an essential mediator of cell polarization and lineage specification in response to asymmetric cell‐matrix adhesion. This study highlights the utility of precisely controlling 3D ligand presentation around single cells to direct cell polarity for regenerative engineering and medicine.
A primary cilium, made of nine microtubule doublets enclosed in a cilium membrane, is a mechanosensing organelle that bends under an external mechanical load and sends an intracellular signal through transmembrane proteins activated by cilium bending. The nine microtubule doublets are the main load-bearing structural component, while the transmembrane proteins on the cilium membrane are the main sensing component. No distinction was made between these two components in all existing models, where the stress calculated from the structural component (nine microtubule doublets) was used to explain the sensing location, which may be totally misleading. For the first time, we developed a microstructure-based primary cilium model by considering these two components separately. First, we refined the analytical solution of bending an orthotropic cylindrical shell for individual microtubule, and obtained excellent agreement between finite element simulations and the theoretical predictions of a microtubule bending as a validation of the structural component in the model. Second, by integrating the cilium membrane with nine microtubule doublets, we found that the microtubule doublets may twist significantly as the whole cilium bends. Third, besides being cilium-length-dependent, we found the mechanical properties of the cilium are also highly deformation-dependent. More important, we found that the cilium membrane near the base is not under pure in-plane tension or compression as previously thought, but has significant local bending stress. This challenges the traditional model of cilium mechanosensing, indicating that transmembrane proteins may be activated more by membrane curvature than membrane stretching. Finally, we incorporated imaging data of primary cilia into our microstructure-based cilium model, and found that comparing to the ideal model with uniform microtubule length, the imaging-informed model shows the nine microtubule doublets interact more evenly with the cilium membrane, and their contact locations can cause even higher bending curvature in the cilium membrane than near the base.
The nuclear lamina is widely recognized as the most crucial component in providing mechanical stability to the nucleus. However, modeling the diverse mechanical aspects of this protein network still poses a significant challenge. We developed a constitutive model for the nuclear lamina network based on its microstructure, which accounts for the deformation phases at the dimer level, as well as the orientational arrangement and density of lamin filaments. Instead of relying on homology structure, we conducted molecular simulations to record the force-extension response of a highly accurate lamin dimer structure obtained through X-ray diffraction crystallography experimentation. Furthermore, we devised a semi-flexible worm-like chain extension-force model as a substitute for the multilinear equation, incorporating the stretching of the backbone, uncoiling of the dimer coiled-coil, and unfolding of alpha helices. Subsequently, we implemented our extension-force equation within a 2D network continuum model for the nuclear lamina and derived stress resultants. This enabled us to integrate the nuclear lamina into finite element simulations of the entire nucleus. Finally, we conducted simulations of nucleus endothelial transmigration, investigating the impact of varying network density and uncoiling constants on the critical force required for successful transmigration. The model allows us to incorporate the microstructure characteristics of the nuclear lamina into nucleus-related finite element modeling, thereby gaining insights into how laminopathies and mutations affect nuclear mechanics.
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